Special Topic of Forensic Science
Thin Layer Chromatography (TLC)
A type of chromatography that uses silica gel or alumina on a card as the medium for the stationary phase Advantages: Ability to run several samples simultaneously Apparatus is simple and cheap Flexible Sample will remain where dried Dried sample not prone to detection problems Disadvantages: Lower sensitivity and resolution than GC and HPLC
Gas Chromatography (GC)
A type of chromatography used to separate vaporizable compounds; the stationary phase is a crushed metal or polymer and the mobile phase is a nonreactive gas. Disadvantages: samples must be volatalized and derivatized to mitigate deterioration
If you inherit type A from your mother and type O from your father, what is your resulting GENOTYPE? A. AO B. AA C. A D. O
A. AO
Which area would be more susceptible to degradation on the sample item: A. Area in the center of a heavy blood stain B. Area along the edge of the heavy blood stain C. area far removed from the large stain
A. Area in the center of the heavy blood stain
Which vessel is under the most pressure in the circulatory system? A. Artery B. vein C. capillary
A. Artery
Which types of biological samples should never be stored in plastic containers for more than two hours? A. Heavily soaked blood stained items B. Bone fragments C. Teeth D. Stomach contents E. All of the above
A. Heavily soaked blood stained items
A vaginal swab that test positive for acid phosphatase, but shows no spermatozoa on the microscopic examination should be tested further for the presence of what protein? A. P30 B. P50 C. Flavins D. Oligospermia
A. P30
What are the ABO genotypes?
AA, AB, AO, BO, OO, BB
what is the most commonly used confirmatory test for blood?
ABA Card hematrace
the fingerprint database is called:
AFIS
What're the two most popular presumptive tests for semen?
AP spot and BCIP
Evidence examination, DNA extraction and PCR set-up [are/are not] conducted in separate places or at separate times.
ARE (DAB standard 6)
This test works fine with older blood samples. Titerd antiserum is added to the stain. Not very sensitive and lots of blood needed.
Absorption Inhibition Test
Range of Standard Curve and Control Samples
Acceptable range of the assay--derived from linearity studies and depends on application of the procedure. The analytical procedure should provide acceptable linearity, accuracy and precision. Standard Curve--constructed so that max calibrator is 120% of the expected maximum conc and the min is at least 10% than the min quantifiable value. The min quantifiable value is supported by chromatographic data and depends on the degree of background noise.
What are the roles of an expert witness?
Establish credibility through credentials, background experience. Evaluate evidence. Render an opinion about the evidence.
T/F It is the analyst's responsibility to give the investigator opinion on the case and results.
False, the analyst should report finding in a succinct/unbiased fashion, keep thorough documentation of handling/testing, maintain COC and examine evidence to locate/isolate/identify items of serological value
T/F DNA's base pairs are AG and TC.
False, they are AT and GC
T/F WBC's contain no DNA
False- It is RBC's that don't
T/F Oxygenated blood is purple.
False- it is bright red
T/F Males who are labeled as being oligospermic probably had a vasectomy bc it means there is no sperm in the semen.
False- it only means that there is little sperm
T/F the most common blood phenotype is A
False- it's O
T/F You are a Secretor if L a+b-
False- you must be L a-b+
T/F Sperm can remain intact for years when dried on clothing.
False-but it can for months
T/F Sperm don't lose their tail til they die
False-they lose it right after ejaculation
T/F: MSDS does not contain information regarding storage, transportation & disposal methods of chemicals
False. Also includes composition, health hazards, first aid, handling,flammability,toxicology, physical and chemical properties, and regulatory info.
T/F The larger the caliber, the larger the pellet from the shotgun.
False. The numerical value and the size of pellets are inverse.
T/F The phenolphthalein presumptive test is more sensitive than luminol?
False; Luminol is sensitive but reacts with bleach, copper, and iron as well
T/F bottles should be stored above eye level
False; below eye level
T/F regarding MSDS, chemical manufacturers are required to perform exhaustive studies of health hazards and toxicology
False; they are required to report only what's known about the compound.
Field Samples
Field--defined as a location other than the lab Field Documentation of samples---must be recorded in ink on bound notebook that have numbered pages, info for each sample should include: date and time of collection, name of person collecting sample, ID#, documentation that sample collection materials are clean, secure sample under specified conditions Collection of Forensic Samples (ex. urine)--consider the privacy of the individual and the integrity of the sample 1. specified toilet facility used 2. add a colored cleaning agent to the toilet, prevents dilution of sample 3. remove running water and soap from the toilet facility 4. receive specimen immediately 5. check specimen temp, seal ctgr, personnel and subject should initial the ctgr
What profession focuses on criminal and civil investigations of patient traumatic injury or treatment, including both victims and perpetrators?
Forensic Nursing
What profession is associated with this logo?
Forensic Nursing
Instrument Qualifications
--Consider the required level of precision and accuracy prior to purchase --Think about the working enviro (electrical, plumbing) and space needed --Technical and service support provided? --Price-new and replacement parts --# and size of samples, speed of analysis --installation? --operation? installed by vendor? --how many units of this instrument will we need? --special considerations (temp, DNA reagents should not be stored in freezer that has auto thaw)
Validation of Analytical Procedures: Accuracy and Precision
Accuracy-- can be verified by comparison of the results of the proposed analytical procedure with those of a second well-characterized procedure. should be done using at least 9 determinations with three conc levels. Generally reported as the diff bt the mean and the accepted true value together with the confidence intervals. Precision--replicate analysis of 3 with varied conc. control samples are performed on the same day over a minimum of two weeks
Describe the process of using hemastix to test a sample for blood
Add a drop of water to strip then rub against stain in question. They contain TMB which is a reagent used independently like kastle meyer.
Describe the method used to test a sample for blood using the Kastle-Meyer or Leucomalachite Green reagent
Add drop of reagent to a stain extract (dry rub on filter paper). Add a drop of hydrogen peroxide and color change is observed. 20 seconds or less = positive. Before hydr peroxide or longer than 20-30 seconds considered neg.
+ rapid and simple + Sophisticated equipment is not necessary + minimal training + minimal sample
Advantages of Color Tests
+ Selectivity - Extraction processes can be designed to selectively extract an analyte or class of analytes. + Reproducibility - SPE eliminates emulsion formation; reduces the dependence on operator technique; improves extraction reproducibility. + Speed - Vacuum manifolds allow the simultaneous extraction of multiple samples; low volumes of solvent employed reduce the time for evaporation steps; SPE techniques are easily automated unlike liquid extractions
Advantages of Solid Phase Extraction over Liquid-Liquid
+ Ability to run several samples simultaneously + Apparatus and the TLC equipment is cheap and simple + Technique offers great flexibility in the choice of mobile and stationary phases + Any sample dried on the plate will remain there + Dried sample is not prone to detection problems associated with non-elution, thermal instability and masking by the solvent front
Advantages of TLC
+ Sensitive and selective + Usually take place spontaneously and rapidly
Advantages of microcrystal tests
What fixative protein-dye is useful for bloody impressions and stains residues a blue-black color?
Amido black
no specific arrangement and melts over a range of 10 or more degrees. melt at lower temperatures than pure crystalline substances.
Amorphous Form of a drug
Describe the way the phadebus test works
Amylase substrate is in an insoluble blue dye, linked to starch. Amylase in sample hydrolyses polymer; can be measured colorimetrically at 620 nm Absorbance in value that exceeds 0.3 units considered positive.
postmortem investigations, workplace drug testing, and human performance investigation.
Applications-Immunoassays have applications in
List ONE of the three general fingerprint pattern types.
Arch, Whorl, Loop
What are examples of mineral fibers?
Asbestos, rock wool, fiberglass
Key Parameters to consider when validating a process
-purity an identity of the test sample -homogenity of soltns -extraction recovery -range of std curve and control samples -linearity and precision of std curves -sensitivity -specificity -accuracy and precision -stability -quality control for DNA validation should be divided into developmental and internal validation
why should you not use Vd to calculate amount ingested
Assumptions made: drug is in equilibrium, dose is fully absorbed, linear kinetics, chronic v. acute users, no elimination has occurred
Which part of the spermatozoon contains DNA? A. Tail B. Head C Both D. Neither
B. Head
Which of the following statements is incorrect regarding Luminol? A. Luminol requires complete darkness B. Luminol is the agent of choice for detecting the presence of blood stains C. Luminol is considered to be a presumptive test for blood D. Luminol is best used for locating the presence of hidden blood stains
B. Luminol is the agent of choice for detecting the presence of blood stains
On which type of surface would you expect more spatter to occur? A. Smooth B. Rough C. They're the same
B. Rough
Which one of the following is an example of wear evidence? A. The handle of the weapon B. Skin cells on a shirt collar C. Both D. None
B. Skin cells on a shirt collar
Hemoglobin derivatives exhibit a strong absorbance at wavelength 400-425nm. This is known as the A. Sorbet band B. Soret band C. Sorrel band D. Squirrel band
B. Soret Band
What component of blood becomes activated when the circulatory system is breaded and loss of blood occurs? A. Hemoglobin B. Platelets C. Glucose and hormones D. Fibrin
B. platelets
This 1979 policy expanded the scope of quality management from military contractors to include general quality requirements for industrial, commercial and governmental products and services
BS 5750 policy (British)
Silyating Agents for GC (used for derivatizing drugs)
BSTFA or BSA are the most widely used derivatizing agents. Compounds that ctg carboxylic acid, amide, phenolic, or OH groups react well with silyating agents. They are highly volatile and readily hydrolyzed back to the parent compound. This means that they should be kept away from water.
A Questioned Document Examiner may be employed by a government agency or what other private type of business?
Banks
Dillie-Kopanyi Test
Barbiturates - Purple
How do you decide what pattern to use?
Based on size/location of crime scene and number of investigators.
What is refraction?
Bending of light as it moves from one medium to another Example- air to water
What type of distribution can be used when an experiment has 2 possible outcomes?
Binomial Distribution
Regarding NFPA hazard code symbols: what do the colors blue,yellow, red and white mean?
Blue: signifies health Red: Flammability Yellow: Reactivity/dangers White: special warnings
State four of the ways that help to estimate time of death
Body temp, Rigor mortis, dulling of cornea, insect infestation
An acid is a substance, charged or uncharged, capable of donating a proton: a base is a substance charged or uncharged, capable of accepting a proton from an acid. • Acids and bases can be either neutral compounds or charged ions • An acidic substance can only behave as an acid in the presence of a base as defined by the Bronsted Lowry theory • Similarly a basic substance can only behave as a base when in the presence of an acid
Bronsted-Lowry Theory of Acids and Bases (Acid Base Equilibria)
What is the name of the document which maintains and helps prove evidence integrity was maintained?
Chain of Custody
A buffer is then passed through the extraction cartridge to introduce a pH environment conducive to the retention of the analyte of interest on to the column
Change of pH Environment in (TLC)
What is the purpose of EDTA during the PCR reaction?
Chelator
Which extraction method is considered too harsh for old or degraded DNA?
Chelex
Type of DNA extraction that uses a resin that has an affinity for metal ions. Addition of heat causes the DNA to break down.
Chelex Extraction
can result in loss of pure drug by breakdown to another compound Example: cocaine to the metabolite benzoyleconine).
Chemical Degradation
What is NOT a part of the ABI 310 Genetic Analyzer? Capillary, CCD camera Syringe with polymer, Chiller
Chiller
what type of stain is commonly used in the US for sperm?
Christmas Tree stain
What type of biohazard hoods protect both the operator and sample when dealing with DNA?
Class II biohazard cabinets
List a type of chemical examination for controlled substances.
Color Chemical Test
tests involve the reaction of a drug sample, a protein free filtrate or an extract with a reagent or a series of reagents to produce a color or change in color.
Color Tests
Oxidation or reduction reaction
Color tests are largely reagent tests and involve a change in color of the reagent by
What type of medullas do humans usually have?
Continuous, fragmented or absent
Quality Control
Control samples are injected at least every 10 samples and each control is injected a minimum of 3 times. They are assessed after each assay run to ensure suitable within batch stability and reproducibility. Specimens from a given animal should be assayed in the same batch.
What substances should be stored under lock and key with their use documented?
Controlled substances
Which layer has pigment granules?
Cortex
What are examples of cellulose fibers?
Cotton, rayon, cullulose acetate
What are some examples of natural fibers?
Cotton, wool, silk, hemp
Which crimes do we place more investigative emphasis on?
Crimes against people
extent to which other drugs and metabolites interfere may give false positive assay results (the test indicates the presence of a specific analyte when it's not actually present, instead it has cross -reacted with a structurally similar molecule).
Cross Reactivity
the test substance has dispersed or dissolved in the test reagent, and the mixture becomes saturated,
Crystal formation occurs when
term used in screening assays. Such assays identify samples as either positive or negative but provide no quantitative results or drug concentration. A 'cut off' is the point below which all samples are considered to be negative.
Cut Off Level
Which layer is resistant to chemical decomposition?
Cuticle
What are the 3 layers of hair?
Cuticle, cortex, medulla
Which one of the following statements is true in relation to DNA: A. Adenine is a pyrimidine base B. Cytosine pairs with Adenine C. DNA forms a triple helix D. Adenine binds to Thymine with two hydrogen bonds
D. A-T bind by two hydrogen bonds; G-C bind by three. Adenine is a purine base and Cytosine pairs with Guanine
When using an orange filter on an alternative light source, which biological fluid will fluoresce white? A. Vaginal secretions B. Semen C. Urine D. All of the above
D. All
Which of the following reagents is a protein stain? A. Ponceau red B. Coomassie blue C. Amido black D. All
D. All
The cellulose based paper, used for the storage of whole blood, that prevents DNA degradation and microbial breakdown is known as A. EDTA paper B. STR paper C. Nucleic Acid paper D. FTA paper
D. FTA paper
Which male gland produces PSA? A. Thymus B. Seminal Vesicle C. Thyroid D. Prostate
D. Prostate
Which of the following is not true regarding RFLP analysis: A. DNA fragments are cut with restriction enzymes B. RFLP requires the use of n blotting C. DNA segments are separated using gel electrophoresis D. RFLP uses less sample than current STR analysis
D. RFLP uses Less sample than STR (it uses more)
Which is NOT considered one of the most important aspects of crime scene investigation? A. Securing the scene B. The prevention of contamination C. The preservation of physical evidence D. The protection of Onlookers
D. The protection of onlookers
Which of the following is NOT a typical search pattern? A. Strip or Lane search B. Grid search C. Spiral search D. Zig Zag search E. Quadrant search
D. Zig Zag search
Electronic Records-Life expectancy
Data Storage Type Life Expectancy Magnetic tape 5-20 yrs CD-ROM 2-100 yrs Paper 20-100 yrs Microfilm 10-200 yrs
A pH value is a number that is a measure of the acidity of a solution. It is defined by the equation pH = -log10 [H+] This is not strictly correct as a single proton does not exist in aqueous solution but is hydrated: H+ + H2O --> H3O+ (hydronium ion) so pH = -log10 [H3O+] (H+ and H3O+ may be used interchangeably)
Definition of pH
Schedule
1: High abuse, no medical, no safety 2: High abuse, medical use, high addiction 3: Less high abuse, medical use, moderate addiction 4: Lower abuse, medical use, limited addiction 5: Low abuse, medical use, little addiction
How many air changes per hour will provide adequate air quality in the lab?
4 to 12
# of calibrators needed to construct a standard curve
4-7 but depends on the range of concentrations being quantified, also a min of 3 control samples at low, intermediate and high conc values should be prepared separate from the analyst performing the assay procedure. High control sample = 10% less than the max std curve calibrator Low control sample = 10% greater than the min std curve calibrator
When taking reference samples from the head, how many hairs do you need?
50 full length hairs
Describe how the ABA Card Hematrace works, and state one reason why investigators may not use it
Detects Hgb. Based on antibody-antigen rxn to antibodies in test card. Similar to preg test. Positive rxn w/ ferret blood - some labs will not use and some will process but give this disclaimer
Psychology/Psychiatry
Determine competency of a suspect to stand trial
Anthropology
Determine if bones are human or animal
Odontology
Determine skeletal remains through dental records
Which extraction methods is most appropriated for a vaginal swab from a sexual assault kit?
Differential
What is the main type of buffer used in organic extraction?
Digest buffer (contains SDS and pro-K)
wetted to provide an environment suitable for the reproducible retention of the isolate. This is achieved by passing several bed volumes of a suitable solvent such as methanol through the column.
During the extraction sequence in Thin Layer Chromatography, the sorbent must first be
Typical data applied to an electropherogram does NOT include A. name of locus B. number of repeats C. size of the fragment D. peak height E. peak area
E. Peak area
drug is labeled with an enzyme and the measurement is an optically detected change such as color change, ultraviolet absorption, fluorescence or luminescence. In these types of immunoassay there is a difference between the optical signals generated by the free and bound labeled drug, so no separation steps are required before analytical measurements are made Homogenous Assay
ELISA - In enzyme linked immunosorbent assays
a homogeneous, liquid phase assay. It relies on the ability of free antibody to bind to enzyme-labeled drug and inactivate the enzyme. This requires that (1) enzyme linked to the drug must retain its activity, and (2) the binding of enzyme-labeled drug to the antibody inactivates or severely reduces the activity of the enzyme producing a color change
EMIT - Enzyme multiplied immunoassay technique
what presumptive test for feces has been disputed but reacts with urobilinogen?
Edelman test
Opioids
Effects: Analgesia, euphoria, cough suppression, mitigate diarrhea, miosis From: opium plant, or produced synthetically Used: intravenously, inhalation, orally, transdermally Actions: Modulates membrane receptors, inhibits neuronal transmission, inhibits adenylyl cyclase Death: respiratory depression from high doses, suffocation from vomiting
Volatile Substances
Effects: CNS depressants at high doses, may cause stimulation at low doses, ataxia From: cans and synthetic materials Used: inhaled Actions: Stimulate GABA receptors, inhibit NMDA receptors mediate CNS depression Death: Direct effect on CNS, cardiovascular or pulmonary toxicity or indirectly from vomit
Benzodiazepines
Effects: CNS depression, PNS relaxation, minor suppression of cardiovascular From: Synthetic, benzene and diazepine ring Used: Orally Actions: Binding GABA post-synaptic receptor, do not open Cl- channels Death: Unlikely, insomnia most likely
Barbiturates
Effects: CNS depression, suppression of PNS and cardiovascular From: Synthetic from urea, malonic acid Used: orally Actions: Post-synaptic GABA receptor, increase Cl- flow Death: Lethal depressant effects on respiratory and vasomotor centers
Alcohol
Effects: Vasodilation, depresses contractility, depression of CNS, can irritate gastric system From: Sugars converted Used: Orally, rectally Actions: Increase inhibitory effects of GABA, stimulate activity of dopamine system, increases serotonin levels Death: From respiratory arrest BAC +0.45
Hallucinogens
Effects: euphoria and hallucinations From: organic and synthetic sources Used: orally, ingested, inhaled Actions: serotonin agonist, can have dopaminergic agonist, cholinergic actions, norepineprhine like, acetylcholine like Death: Very unlikely
Cannabinoids
Effects: euphoria, treatment of nausea and vomiting, stimulate appetite, analgesia From: extracted from cannabis plant Used: Ingested, inhaled Actions: bind to cannabinoid receptors, stimulate intracellular cascades Death: Unlikely, from prolonged smoke exposure
CNS Stimulants
Effects: increased cardiovascular actions, CNS stimulation, euphoria, From: synthetic and natural sources Used: orally, inhaled, ingested, insufflated Actions: can block nerve channels, indirectly acting sympathomimemtic drug, potentiates monoamine neurotransmission such as norepinephrine, dopamine or serotonin Death:
Hallucinogens Final Color, Violet
Ehrlichs reagent
Which step of DNA amplification involves the use of DNA (TAQ) polymerase? Denaturation, Primer annealing, Elongation, or termination
Elongation
Finally, the analyte of interest is eluted from the column and collected into a tube using a suitable solvent. The strength of the eluting solvent can be adjusted to optimize the elution of the analyte of interest while at the same time minimizing the amount of endogenous material removed from the column.
Elution (TLC)
there is another substance present to accept it. This may be another dissolved substance or the solvent itself
A proton can only be donated when
What color do MDMA-type drugs turn under marquis reagent?
*blue to black*
what color does amphetamine & methamphetamine turn under marquis reagent?
*orange*
what color do opium alkaoids turn under marquis reagent?
*purple*
Work place records
There are 3 types of work place records: Records--can document or facilitate the functioning of a lab and can include policicies, decisions, info mandated by federal or state law or by granting agencies, minutes of meeting and telephone logs Non-records--documents that do not further the understanding of the work in the laboratory or the program. They may be copies of an original record that is archived Personal Papers--papers concerned with an indiv. personal business ex. books or similar items purchased with personal funds
What is the function of the RBC's?
Tissue oxygenation
T/F CODIS looks for relationships between the profiles of unidentified remains and relatives of missing persons.
True
T/F Carbonation enhances the abs of OH-
True
T/F Chain of custody is required for any evidence submitted to court and can be used to establish the integrity of the evidence?
True
T/F DNA is a negative molecule, is an acid, is carried in the nucleus of cells, it's main components are sugar-phosphate
True
T/F DNA is double stranded
True
T/F Decomposition of a body in water is slower. This is due to the fact that the water temp is cooler.
True
T/F Fingerprints are individual characteristics
True
T/F Fingerprints remain unchanged for a person's lifetime
True
T/F Glass breaks 1st of the opposing side of the force
True
T/F Handwriting analysis can provide individual and class characteristics.
True
T/F Human sperm is larger than bacteria and yeast.
True
T/F Human sperm is smaller than dog sperm or skin cells.
True
T/F Hydrolyzed body fat can help preserve a body and prevent putrefaction?
True
T/F If the angle of impact is = to 90 then the blood drop is circular.
True
T/F In the absence of a hair root, mtDNA testing may be carried out on the hair shaft.
True
T/F International Organization of Standards (ISO) guides are becoming more prevalent as the recognized standard for quality assurance programs
True
T/F LIMS data must be secure and closed, with restricted access to information
True
T/F Mature RBC's do not contain DNA.
True
T/F Mitochondrial DNA analysis may be conducted on hair shafts that contain no root material.
True
T/F Multiplex PCR uses more than one set of PCR primers to target multiple locations.
True
T/F PGM can be found in blood, vaginal secretions and semen?
True
T/F RBC's are 45% of the total blood volume
True
T/F Saliva can be difficult to visualize on ALS because it may not fluoresce as well as other fluids
True
T/F Shotguns have no rifling and shoot several projectiles.
True
T/F Sperm DNA is in the head only
True
T/F Standardcurve and control samples are analyzed w/ each assay batch of samples
True
T/F The WBC's are produced as a response to infection in the body.
True
T/F The amylase activity in saliva is variable from person to person often resulting in inconclusive testing for its presence?
True
T/F The liver metabolizes 90% of OH-
True
T/F The slot blot method of DNA quantitation can measure both double and single stranded DNA?
True
T/F The surface tension of blood is less than that of water.
True
T/F Vaginal fluids, breast milk may produce false positives for saliva / salivary amalyse
True
T/F Video recroding is an acceptable method of recording a crime scene.
True
T/F When inflicting a deep stab wound, the body tissue may "wipe" blood off the weapon as it is withdrawn from the wound.
True
T/F You should wash and dry your hands before being inked.
True
T/F a written SOP should be avail.for every analysis conducted ina lab
True
T/F each analysis for a sample must be established, documented and kept secure
True
T/F informed consent for all human participants in a study is required
True
T/F mDNA is more abundant than nuclear DNA in the body
True
T/F mtDNA is inherited strictly from a persons mother.
True
T/F mtDNA testing is also the method of choice for badly degraded samples such as old skeletal remains.
True
T/F one way of confirming blood is to visualize RBCs under a microscope, but the success of this method depends on the skill of the analyst and age of the stain as well as nature of the surface from which the stain came
True
T/F passive blood deposits are transfer, flow patterns, drips
True
T/F separate areas exist for pre- and post-amplified DNA
True
T/F system suitability is based on the concept that the equipment, electronics, analytical operations, and samples to be analyzed constitute an integral system that can be evaluated as such
True
T/F the CFR mandates that studies funded by the FDA or EPA be conducted using protocols
True
T/F the original chainof custody is included with a sample if it is to be transferred to anotherlabfacility
True
T/F the surface tension of blood is less than water
True
T/F there are no confirmatory tests for urine. presence of urine is only inferred from the positive presumptive tests
True
T/F urine can be detected using ALS and will chemiluminesce a blue-white
True
T/F well maintained instruments usually have better sensitivity and gives more reliable reproducible results than those that are not well maintained
True
T/F: When using an alternative light source to search for biological fluids, the wavelength of light used must be opposite to that of the color of the stain?
True
T/F: a smal semen stain that dries quickly can have intact sperm for months possibly
True
T/F: if the lab personnel has any exposure to human blood, blood components, products made from human blood or other infectious material, annual bloodborne pathogen exposure control program must be implemented.
True
The concentration of OH- is greater in the spinal fluid, virtous humor and saliva than in the blood
True
T/F Access to the laboratory should be controlled and limited.
True (DAB standard 6)
T/F the care and use of animals is regulated
True - 9 FR Animals & Animal Products
T/F even with a published method, a lab should still write its own SOP for an analysis. why??
True - in-house will be more detailed and eliminate possible inconsistencies in the results
T/F personnel should enter their facility through a checkpoint or use a card key to ensure integrity of samples and results
True.
T/F An IACUC must review and approve all studies involving animals
True; IACUC = institutional animal care and use committee
T/F MSDS are internet accessible and can be used in lieu of a hardcopy
True; its advisable to have hardcopy MSDSs
Decide which statistical test is associated with the info for each: This is a post-hoc test that is used for testing the significance of unplanned pairwise comparisons
Tukey's Honest Significant Difference test
a swipe bloodstain pattern is the result of
a bloody object in motion contacting another clean surface
disadvantages to microcrystal tests
impurities can interfere with crystal formation; training and expertise needed to ID drugs; some degree of subjectivity involved
(accuracy, precision) is when the analytical procedure is used to assay a reference standard of known concentration
accuracy
white five factors must be demonstrated before an analysis is used, regardless of SOP?
accuracy, precision, selectivity, reproducibility and repeatability of a method
weak acid because it does not readily dissociate
acetic acid
Most presumptive semen tests look for the presence of what substance found in high quantities in semen?
acid phosphatase
screening tests for semen are based on detecting the enzyme
acid phosphatase
Solvent effects
acids can only donate protons when the solvents can accept protons
agonists
activate the receptor, cause pharmacological activity, bind as substrates to receptor sites, promotes response
Acetylating agents for GC (used for derivatizing drugs)
agents such as acetic anhydride, propionic anhydride and trifluoroacetic anhydrides are the most commonly used. These agents are more stable than silyl dertivatives and are useful with many classes of drugs.
What is the protein that helps to maintain blood volume?
albumin
the change in body temperature post mortem is called
algor mortis
Groups that do NOT significantly ionize in water
alkanes, alkenes, alkynes, ketones, amides, ethers
Controlled Substance Storage
all controlled substances should be stored under lock and key and their use must be documented, most common is ethanol
double antibody technique
soluble drug-antibody complex is rendered insoluble by adding a second antibody (ppt antibody targeted to the globulins (albumin) of the animal in which the first antibody was raised. Separation can then occur through centrifugation, the heavier double antibody complex forming a pellet that can be recovered and measured
why should plumbing be made of acid-resistant materials?
some chemicals will undoubtedly be poured down the sink
Drugs that exhibit Protein Binding
some drugs bind to proteins, to remove the drug from the proteins an analyst must heat the sample in acid or incubate with proteinase enzyme
The "Christmas Tree Stain" is a confirmatory test for
sperm
who approves and signs the study protocol?
sponsor and study director; QA officer and consultants can beinclued
toxicologist
studies body fluid, tissue, and organs for drugs and/or poisons; may conduct postmortem pathological examinations, and examination of personal effects and empty containers, etc.; may have extremely minute quantities to test; must detect, identify, quantify, and assess toxicity
volume of distribution
the theoretical volume of body fluids in which the amount of drug in the body is distributed at a concentration equal to that of plasma. highly polar/ acidic drugs have low Vd, basic drugs have a high Vd. assume distribution and equilibrium have already occurred. based on steady state and plasma, not blood, should not be performed postmortem
up to 24 hours
the time that drugs remain in the bloodstream
up to 72 hours
the time that drugs remain in urine
distribution
the transport of absorbed drug by the bloodstream to all parts of the body
who can be held legally responsible for accuracy of reported results?
the manager. employees should have access not only to his/her immediate supervisor but also other members of management.
Mass Spectrometry
the sample is in vapor form and is bombarded with e-, which converts the substance to an ionic state which consists of the parent ion and several ionic fragments of the original molecule. They are separated according to their mass/charge ratio. The MW of the compound can be deter from it's spectrum. ADV. --specific- best analytical technique, sensitive ng/ml, qual and quant, can be used with separation techniques DIS--expensive, high maitenence, trained analysts, less sensitive than Fluorescence, best for low MW < 500, can be issues LC
Solvent effects in UV-Vis
aromatic chromophores show fine vibrational in non-polar solvents, in aq the pH affects ionizable chromophores due to conjugation changes in the ionized and non-ionized chromophore bathochromic shift--in phenolic compounds when the soltn in more alkaline it leads to an abs shift to a longer wavelength (red) ex. barbiturates hyerchoromic effect--a loss of fine structure and an increase in molar absorbtivity
Immunoassay--Cut Off Levels
assays ID samples as either positive or negative but provide no quantitative results or drug concentration. So the cut off is the point below which all samples are considered to be negative. Can also act as the limit of detection
what process helps ensure the laboratory is maintaining accreditation standards?
audits
collected writing samples should be
authenticated and from an appropriate time period.
Final Report: Research Plan
background info and study objectives are included along with experimental design and methods. FDA and EPA require--description of test articles, description of test system, description of methods used
what class does Dillie-Kopanyi test for and what is the color associated with it?
barbituates; *purple*
describe how the AP spot test works
based on hydrolysis of phosphate esters; acid phosphatase reacts w/ sodium-alpha-napthylphosphate and fast blue B to produce purple/blue color. Positive result is intense color in 20-30 seconds. can be 1 step or 2 step process. Can use AP reagent on filter paper rubbed from stain.
immunoassay
based upon specific drug-antibody reactions; the best method for detecting low drug levels
Is the pH of saliva acidic or basic?
basic
the strength of an acid depends not only on its ability to give up a proton but also on the ability of the solvent to accept it.
basic strength of the solvent.
Sample condition description
be sure to include features and appearances of the substance that will be tested such as: --weight (dry or wet) --amount if in tablet form --# of packages --nature of substance --Color --odor --condition (wet or dry)
Why is CO toxic to humans?
because the CO binds to hemoglobin and then the oxygen can't
Schedule IV
benzodiazepines, carisoprodol, tramadol
Schedule IIIN
benzphetamine, phendimetrazine, ketamine, anabolic steroids
Pharmacodynamics of Methamphetamine
blocks neurotransmitter reuptake, reverses reuptake transporter, displaces neurotransmitter from vesicular storage sites, all will increase neurotransmitter in the synapse
which is NOT a variable to cinsider when examining blood patterns?
blood alcohol level when blood is shed
directly proportional
blood and brain blood concentrations
What is the cell free part of the blood called?
blood plasma
What is the liquid that remains after blood clots?
blood plasma
screening
broad analyses performed in order to determine what types of drugs are present, various instruments (immunoassay, spectrophotometry, GC, LC)
substrate samples collected from the scene of a suspicious fire should be taken from
burned and unburned areas
blood analysis
calculated with a high degree of accuracy by use of gas chromatography which separates alcohol from any other chemicals
List info you can get from examining the bullet casing
caliber, manufacturer, compare primer impression
difference between calibration and standardization?
calibration = checks physical measurements. standardization = determines a response fxn
what is "a comparison o fa measurement standard,instrument or item with a standard or instrument of higher accuracy to detect and quantify inaccuracies and to report or eliminate those inaccuracies by adjustments"
calibration- EPA definition
subcutaneous
can be performed by patient, more rapid than oral, painful (needles), variable absorption
hair
can have a more permanent marker of drug intake
What 5 effects does heat have on electrophoresis?
can melt agarose gel, break glass plate, denatures and deactivates proteins, distorts band shapes so may produce smiling gels, bands can become thicker
Study Protocol
can refer to: a prelimiary memo by diplomatic negotiators as a basis for a final treaty ex. ozone layer, the formatting and treatment of data in an electronic communication system, a strict code concerning proper etiquette and rules of precedence, a detailed plan of scientific or medical procedures for QA/QC work, a study protocol is a document that lays out the objectives and a detailed description of the total experiment or research activity.
what substances have been known to cause false positives of presumptive semen tests?
carrot/cauliflower juices, blood, male urine, fem hygiene prods, spermicides, vaggie fluids, blood
Solid Phase Extraction
cartridges consist of a polyproylene column ctg bonded silica sorbent material held in place by an upper and lower frit.
what aspects must be included in a forensic court report?
case identifier, description of evidence, description of methodology, locus used, results/conclusions, interpretative statement, date issues, disposition of evidence, signature line
This type of blood spatter can be useful in determining how many blows occurred during an assault
cast-off spatter
absorption of energy
causes molecules to oscillate, inducing electric and magnetic fields
crime scene documentation methods do not include:
cell phone records
Central stockroom
central to all laboratories, makes everything more efficient and cost effective than have separate stock rooms
metabolism
changes to the drug to facilitate its removal from the body, mostly occurring in the liver
4 things necessary in developmental validation:
characterization of the locus, species specificity studies, sensitivity, stability & mixture studies.
what should the technical manager be familiar with?
instrumentation, calibration or test methods, procedures, assessment of results
what are possible causes for statistical outliers?
contamination of sample, instrument malfxn, technician error, over-titration
UV or Vis Components: Light Source
continuum light source is used, power does not change over a wide range of wavelengths, UV range = 160-175nm, typically H or Deuterium lamps, Vis range = 350-2000nm, Tungsten (W) lamps are most common
transdermal
convenient, avoid drastic fluctuations in dose, good for low dose/high potency, delayed onset, prolonged duration of effects, skin is a good barrier (variable absorption)
what are 3 characteristics of the cannabis sativa plant, with only 1 being specific?
covering hairs, cystolithic hairs, glandular hairs
Three specific cellular features of cannabis sativa
covering of cystolithic hairs (not specific), glandular trichomes (distinguish plant as cannabis sativa-specific), covering hairs (not specific)
What two major components of urine are often assayed?
creatinine and urea
inhibitory aa neurotransmitters
decrease the excitability of target neurons, activate receptors that hyperpolarize the post-synaptic membrane
Why there should be an area separate from holding unanalyzed specimens in which any specimen can be stored under appropriate conditions
development of new technology and future analysis; and further analysis for court proceedings/external agencies.
what're the two types of validation procedures for DNA labs?
developmental, internal
what are examples of sample preparation procedures that can impact its analytical results?
dilutions, measurement of pH & weight, duration of time during extractions, amplifications, etc.
3 things/places chemicals should not be exposed to when storing:
direct sunlight, another heat source, under a sink
what are advantages to using microcrystal tests for drug ID?
directness, simplicity, sensitivity, rapidity and ability to conserve evidence
Prep for External Inspections: Laboratory Clean-up
discard expired chemicals and stock soltns, remove equip and chemicals that are not used, est that equip and instruments needed are available, check equip files to be sure they are all up to date, check for labels on instruments (ID#, install date, date of last maitenance, etc), check that all bottles are labeled, place samples analyzed, reagent bottles in their proper locations, discard samples and records no longer needed
Give an example of a discrete quantitative variable and a continuous quantitative variable
discrete--means it can be counted, example how many bedrooms a house has continuous--could be a measurement or a decimal, weight from balance, height
The mechanism by which an acid transfers a proton to a base Acids and bases are crudely classified as strong or weak depending on the completeness of the ionization process (extent of ionization).
dissociation or ionization
gunshot residue:
does not travel a great distance
postmortem
done, occurring, or collected after death
common fingerprint minutiae include
dots, bifurcations, islands
Assay Techniques: ELISA
drug is labeled with an enzyme and the measurement is an optically detected change such as color change, UV abs, fluorescence or luminescence. There is a diff bt the optical signals generated by free and bound labeled drug, so no separation steps are required before analytical measurements are made. This is a homogeneous assay bc it does not require the separation step. Disadvantages-less , only measure drugs to ng/ml, this is why IRA is the method of choice
hair
drugs can remain permanently in _____ protein
enterohepatic circulation
drugs may be excreted from liver to bile, which may then be reabsorbed in small intestine (GI tract) or excreted in feces. basic drugs can passively diffuse across GI mucosa from blood
24
drugs remain in the bloodstream for ____ hours
72
drugs remain in the urine for _____ hours
Fluorescence spectrophotometry
emission of light Advantages: increased sensitivity, increased specificity Disadvantage: few drugs naturally fluoresce, inability to distinguish between parent and metabolite
Disadvantages of Liquid-Liquid Extraction
emulsions may form trapping the drug, immiscible solvents are required, time consuming, glassware clean-up, large volumes of solvents used, skill involved by operator
What's the purpose of an IRB w/ human subjectes?
ensuring their safety and confidentiality of all information for each individual.
What is the name of the molecular structure of an antigen that complexes with a binding site on an antibody?
epitope
Amphoteric compounds
exhibit both properties of acids and bases, optimal extraction of these chemicals can only be achieved by adjusting the pH to get the drug into an unionized form. Ex. morphine, pH = 8
Cross-reactivity
extent to which other drugs and metabolites interfere and may give false positive assay results, may be concentraiton dependant Cross-reacting Substances Amphet/Methamp MDA, MDMA, Ephedrine, pseudophedrine, fenfluramine, phenylpropanolamine, phentermine, phenmetrazine Benzodiazepines Chlorpromazine Cocaine (BZE) Ecgonine, ecgonine methyl ester, cocaine THC (cannabinoids) IBU, naproxen, ketoprofen, tolmetin, acetylsalicylic acid (aspirin) LSD Ergotamine, tricyclic-antidepressants, verapmil, sertraline, fentanyl Morphine Codeine, hydrocodeine, thebaine, hydrocodone, dihydromorphine, hydromorphone, oxycodone, oxymorphone, meperdine, norcodeine PCP TCp, Diphenhydramine, Dextromethorphan
Bioavailability (F)
factors that effect a drugs ability to reach the systemic circulation: water soluble, lipid soluble, transporter help, enzyme destroyers, protein protectors?
It is not possible to examine a document to see what printer or copier it could have originated from
false
T/F post-amplified DNA waste can be stored in the pre-amplified DNA area
false
T/F when working w/ bloodborne pathogens, gloves should be reused
false
fingerprints change over the course of a person's lifetime
false
most reconstruction patterns are easily transported to the lab for further analysis
false
pushing a tool into the surface makes a striated tool mark
false
questioned document examiners prefer to work with copies so they do not damage the original document
false
sliding a tool into the surface makes an indented tool mark
false
there is a scientific way to precisely determine the time of death
false
there is a useful confirmatory test for the presence of saliva
false
you should only document what you see
false
T/F subjects fitting stated criteria in a study cannotbe paid
false - can be paid or unpaid volunteers
the synthetic opiate we discussed in class that was unknowingly injected by drug users and caused near instant death was
fentanyl
Describe the process of the hemochromogen (Takayama) test
ferrous iron from Hgb reacts w/ pyridine to produce red feathery crystals. They are observed w/ light microscope.
What is the protein that helps trap platelets and form blood clots?
fibrogen
how can a sample be maintained when collected in thefield and transported to lab?
fill out chain of custody with all info, seal sample containers in zip log bags; seal and lock the shipping container and examine it for signs of tampering
this is not a fundamental principle of fingerprinting
fingerprints are formed when someone is about five years old
Fluorescence Polarization Immunoassay (FPIA)
homogeneous (inversely proportional) - analyte is covalently linked to fluorophore to make a fluorescent tracer, competes with analyte for binding to antibody, if bound to antibody motion of tracer is reduced, increase observed polarization, MORE drug from sample = LESS polarization
Assay Techniques: EMIT
homogeneous liquid phase assay, enzyme multiplied immunoassay, relies on the ability of free antibody to bind to enzyme-labeled drug and inactivate the enzyme. This requires two things: 1) enzyme linked to the drug must retain its activity 2) the binding of enzyme-labeled drug to the antibody inactivates or severely reduces the activity of the enzyme producing a color change
What is the product rule?
Frequency of a multi-locus genotype is the product of each individual loci frequency
What is the random match probability?
Frequency you would expect to see a DNA profile in a given population group
Standard by which the court uses to decide if a process is accepted by the scientific community
Frye Standard
What court case in 1923 set the "general acceptance" of scientific evidence?
Frye vs. United States
gunshot residue (GSR) testing can be difficult to perform because...
GSR particles do not travel far, can be washed off and only last a few hours
The purity of a compound is usually tested using ______ and the identity of a compound it tested using ______.
Gas Chromatograph, Mass Spectroscopy
2 other ways to describe a normal distribution
Gaussian distribution, bell-curve
What is the SI unit for mass?
Grams
in a strand of DNA, cytosine (C) complementary pairs with
Guanine (G)
Schedule II/IIN definition
High potential for abuse; may lead to severe psychological or physical dependence
conducted on about 1 mg of dry sample, aqueous solutions of the sample, solvent solutions of the sample or solvent extracts of the sample, and are conducted in test tubes or on white tiles.
How are Drug Color Tests conducted
a form of chromatography that uses a mobile phase moving across a thin, uniform layer of finely divided stationary phase by capillary action. Commercially available stationary phases are bonded to a glass plate or a plate made of some other supporting material. A mixture of drugs or drug extract is applied to the base of the plate and developed using the mobile phase. The primary use of TLC is as an identification or separation technique.
How does TLC work
is a strong acid in water since it readily gives up its proton
Hydrochloric acid
• Pharmaceutical products that possess an ester or an amide bond are particularly susceptible to hydrolysis • Esters are usually hydrolyzed by acid or base catalysis • Examples of drugs susceptible to hydrolysis include cocaine and procaine
Hydrolysis
What is the difference between hydrophilic and hydrophobic?
Hydrophilic-water absorbing Hydrophobic-water repelling
Facilities
laboratory operations should be removed from each other to prevent contamination, each room should be equippted to adequately handle the function for which it was designed, separation of air exchanges may be necessary, facility must be secure to ensure the integrity of the samples (check-point, key cards, key pads, id badges), guest must be escorted by personnel, exits in case of fire or other emergency should NOT be limited.
the depressed helical grooves in a bullet are called
land impressions
What are the land and groove impressions made on a bullet?
lands are the uncut metal in the barrel and the grooves are the spiral cuts that are made in the barrel, when the bullet is ejected through the barrel it is marked with a mirror image of the lands and grooves from the barrel.
gunshot residue is characterized by the presence of the elements
lead, barium, and antimony
instrumentation laboratory
less sinks and more electrical power needed, ample floor space and bench tops, be sure there is access to all four sides of an instrument
A Medical Examiner may be able to estimate the time of death by evaluating the stage of decomposition. What stage is occurring when minutes after death a discoloration of the skin occurs from the settling of red blood cells after the blood has stopped circulating? (blood settles in areas closest to the ground).
Liver decomposition
Where is the oldest crime laboratory in the United States located?
Los Angeles
Schedule IV definition
Low potential for abuse relative to III
Schedule V definition
Low potential for abuse relative to IV; drugs with limited quantities of certain narcotics
according to Ms. Clark's lecture, what body part is often missing from a body found in the wild?
Lower jaw
What part of the body does livor mortis occur?
Lowest extremities
what test for vaginal secretions is highly subjective; not widely used; when is a positive called?
Lugol's test; when majority of cells stain chocolate brow from yellow color
What is a visualization-enhancing chemical that causes bloodstains to glow in the dark - even trace amounts?
Luminol
What is water-based, may dilute bloodstains, HIGHLY sensitive but used as a last resort; can help visualize bloody impressions
Luminol
What is the function of albumin?
Maintain appropriate osmotic pressure
Methadone, Final Color, Blue
Mandelin reagent
What are synthetic fibers and what would be an example?
Manmade or altered fibers Acetate, rayon, polyester
Occurs in any sample containing water
Microbial contamination
chemical precipitation tests can be used to identify chemical substances in trace or large quantities. The tests produce crystal derivatives from simple molecular addition compounds of the substance tested and a test reagent. Crystals are then observed using a polarizing microscope.
Microcrystal Tests
Polarized Light Microscopy
Microcrystal reaction products are usually examined by
Short tandem repeats are also known as _______. Microsatellites, Microfragments, Microsegments, Microrepeats
Microsatellites
size, form, color and aggregation characteristics.
Microscopic examination can be used to determine
what is the oldest method of identifying fecal matter, and what characteristics identify it as feces?
Microscopy. Undigested residues of food within last 12-24 hours - characteristic fibrous materials found in animal meat, fruit, grain, veggs & fish
What is a misdemeanor?
Minor Crime
If there is no nuclear DNA present, what type do scientists use?
Mitochondrial DNA
Cost-reducing measures
limit accreditation to those methods being used, submitting applications for several procedures at once rather than successive single procedure submissions, combining the addition of newly accredited methods with an inspection visit, submitting accreditation applications by mail for labs with good track record Cost of accreditation = 25% of profit
zero order elimination
linear relationship, elimination is concentration dependent and constant
Absorptivity (a)
liter/g/cm g/L
the settling of blood in dependent areas post mortem is called
livor mortis
This is the term used to describe the position of a gene on a chromosome
locus
What are the three fingerprint types? (list most to least common)
loops, whorls, arches
Y-STR tests are designed to specifically identify
male DNA profiles
polyclonal
many monoclonal antibodies that have different affinities for the antigen
the decomposition of blood in subcutaneous blood vessels causes an artifact called
marbling
An MS gives an output of what ratio?
mass/charge
headspace
matrix is diluted with a solvent, heated in order to separate volatiles from biological mix, collect gas above liquid
in forensic biology, screening tests
may give false positives
homogeneous assay
measurement of labeled antigen without separating bound and free antigens
Assay Techniques: Radioimmunoassay
measurement of radioactivity (most common 125-Iodine), major drawback is that there is no diff in signal bt bound and unbound drug so they have to be separated first. Heterogeneous assay. The amount of radioactivity bound to the antibody is dependent on the amount of unlabeled drug in the original sample. Bound or unbound fraction can be analyzed but the % of radiolabeled drug bound to the antibody is plotted against the concentration of the unlabeled drug.
Law Enforcement Investigation of DUI
observe vehicle in motion and how driver stops the vehicle, interview with driver in car, administer psychomotor tests (SFSTs) and after driver exits the car (one leg stand, walk and turn, and horizontal gaze nystagmus)
distribution
occurs trough the blood stream
Drug Recognition Experts
officers administer a series of physiological and psychomotor tests, does not determine which drugs are present but may narrow down the class
performance enhancing drug testing - where?
olympics, school sports, organized amateur sports, professional sports,
behavior
once the drug is identified, the toxicologist must assess its influence on the _____ of the individual
antagonistic interaction
one drug interferes with other drug's action - cancels out some or all of one drug's effect
potentiation interaction
one drug makes other drug more effective - enhance receptor performance
liquid-liquid extraction
one of the most common methods, separates based on solubility, polarity, and ionization differences between the compound of interest and the matrix
UV or Vis abs have the following components
one or more light sources, wavelength selector, sample ctgr (cuvette), detector, signal processor, readout device
What 3 types of drugs does Marquis reagent screen for?
opium alkaloids, amphetamine/m ethamphetamine; MDMA-type drugs
Gas Chromatography
the separation of analytes is achieved by their partitioning bt an inert gas (the mobile phase) and a liquid stationary phase held in place by a fine capillary column. At a constant temp. and gas flow rate, different compounds will be retained by the stationary phase for different amounts of time. A compound that preferentially partitions into the stationary phase will have a longer retention time than that of a compound that tends to stay in the gaseous mobile phase.
receptors
protein complexes typically found on the plasma membrane that will recognize specific molecules (neurotransmitters, hormones, drugs)
Antibodies
protein molecules of immune response develop in animals for detection of a specific antigen, produced by the body's B cells in response to a foreign body
urine drug testing purposes
public health and safety, employee safety and fitness, accident investigation, military, workplace reasons, probation and parole
insufflation
quick onset, used for local effects (inhaler), drugs of abuse
microscopically determine the presence of specific cellular features (glandular trichomes).
quick way to determine if an unknown herbal material is marijuana
Protolytic or protolysis Rxns
rxns where an acid reacts with a basic solvent or in which base reacts an acidic solvent to form conjugates.
what is needed in case a chemical is spilled on an employee or his/her clothing?
safety shower or drench shower
GC-FID
sample is carried through a column, based on retention time
toxicology
the study of drugs and poisons, and their interactions with or effects on the body
Covering hairs All of plant Not specific Cystolithic hairs All of plant Not specific Glandular hairs Flowering and fruiting tops Specific
Most frequently encountered structures during microscopical examination of cannabis
hydrolysis, oxidation or photochemical changes, due to the ubiquitous nature of water, oxygen and light in our environment.
Most of the chemical instability of drugs can be attributed to
What type of DNA is found at the hair shaft?
MtDNA- Mitochondrial DNA
1. Antibodies - protein molecules of the immune response developed in animals for detection of a specific antigen 2. Antigens - molecules that elicit an immune response and bind with antibodies 3. A molecular label - may be a totally separate molecule from the antigen, or a modification of it 4. Substrate molecules - used in some immunoassays to react with the molecular label and visualize the antibody-antigen pairing
immunoassays are comprised of a few common elements
Validation of Analytical Procedures: ID and purity of test compound
PURITY --this is usually done via GC or HPLC and the analysis of the sample should show the presence of a single major peak with a retention time corresponding to that of previously injected standards IDENTITY--done via MS
Identity and Purity test of test compound
PURITY is tested by HPLC--should show a single major peak with a retention time similar to injected standards IDENTITY is verified via MS-the full mass spectrum can be compared to standards
the basic chromatographic measurement of a substance in TLC. distance the substance travels from the origin divided by the distance the solvent front travels from the origin. The distance traveled by the substance is measured from the center of the spot if the spot is round. For spots with tails the distance traveled should be determined from the center of the densest area.
RF Values
What 'confirmatory' test for saliva is new, based on antibody-antigen reaction, but has known to have false positives with breast milk, fecal matter and vaginal fluids?
RSID - rapid-stain identification of saliva
What are the types of fracture patterns of glass?
Radial and concentric
ER Spectrum
Radiation Type Frequency Wavelength Transition UV 10 15 - 10 17Hz 200-400nm outer electron Vis 4- 7.5 x 10 14 400-800nm outer electron NearIR 10 12 - 10 14 1 um - 3 um molecular vibrations
analytical measurement is of radioactivity (the most common label is 125I). One major disadvantage of this type of assay is there is no difference in signal between bound and unbound drug so they have to be separated before the analytical measurements are made. Heterogenous Assay
Radio Immunoassay (RIA)
What test comes in prepackaged vials, clear solution that changes to yellow with addition of an extract that contains saliva?
SALIgAE test
what unit is numerical data expressed in?
SI- international system of units
Prep for External Inspections: Review of Operations
SOPS, sample prep, laboratory policies, employee records and training, logs of samples received, records of past issues, corrective actions, maitenance records, laboratory notebooks, validation records, variance letters from accreditation body that approve alt test methods, calibration records, COC
Type of DNA analysis that is currently used. Combines PCR and electrophoresis to amplify and discriminate short repeating units.
STR
Which method uses the smallest fragments of DNA? RFLP, PM+DQA1, STR, AMPFLP
STR
The most commonly used DNA analysis method used today is ___.
STR (short tandem repeats)
when is a microscopic analysis of sperm considered positive?
Sample is positive when any number of intact heads and tails are observed on slide. Presence of three or more heads and no visible tails can also be positive in some labs, depending.
Assuming two donors, consider one locus with alleles 12, 13. 14. How many different ALLELIC combinations are possible for one of the individuals in the mixture?
Six (12/13, 12/14; 12/12; 13/13; 13/14; 14/14)
What two variable affect the diffusion coefficient?
Size and shape of the molecule
What information can a footwear impression provide an investigation?
Size the point of entry and how many people were involved.
• Degradation at the tablet surface • True solid degradation The most common situation involves reactions in adsorbed surface moisture resulting in saturated or concentrated drug solutions. The surface solution phase may result from: melting of the drug or other component and /or adsorbed moisture from the atmosphere.
Solid-state decomposition can be divided into two types:
In organic extraction, what is the purpose of the phenol in the Phenol:Cholorform:Isoamyl alcohol reagent?
Solubilize polysaccharides
What is the name of the document issued to require your presence in court?
Subpeona
Archiving Records
these can be used to: establish where samples were at all times (confirms that the sample could not have been possibly been misidentified), shows that ukn persons could not have tampered with the sample, show that analytical procedures for the sample were followed, interpret data independently These records are really important to forensic evidence bc lawyers often challenge the results of scientific tests
Double Beam Spectrophotometers
these have a mirror that splits the beam emerging from the monochromator. one of the beams passes through the ref soltn to a detector and then a second beam passes through the test sample to a matching detector. the two detector outputs are amplified and their ratio is deter electronically before being displayed. They have the following advantages: compensation for the fluctuations in output of the source, compensation for variations in source intensity and wavelength, continuously recording abs and transmittance data
UV or Vis Components: Refraction Grating Monochromators
these have a reflective surface etched with small grooves, when light enters the monochromator it strikes the grating at an angle and is dispersed, unlike the prism monochromator this provides linear wavelength dispersion along the focal plane of light.
Accreditation Process
to do this a laboratory must demonstrate that it fulfills all the requirements est by the appropriate regulatory bodies, this is demonstrated through a laboratory audit that is conducted by an external, independent expert. In addition, proficiency testing for certification of each analyte must also be done. Analysis of an ukn sample done correctly. Ex. EPA lab analzyes CCl4, benzene or styrene via GC-MS
Comparison of assay results of new procedure
to that of a validated method can be used to show specificity. Ex. peak purity tests using chromatographic or spectroscopic tech may be useful to show that the analyte peak is not attributable to more than one component.
which type of test is frequently done with recovered firearms to evaluate the possibility of accidental discharge?
trigger pull test
T/F a few micrograms of radioactive material not visible to the human eye may be highly radioactive
true
T/F final disposal of samples mustbedocumented
true
T/F: standard curve calibrators and control samples are prepared for each compound of interest (drug or metabolites) ineachbiological matrix to be analyzed
true
T/f microcrystal tests are generally more specific than color tests
true
T/f one reliable confirmatory test for blood is spectroscopy, and has been useful especially in aged and degraded stains that tested negative for color and microchemical tests
true
what statistical test can be used to determine the confidence interval for both small and large populations?
tstudents t-test.
UV or Vis Components:Prism Monochromators
two faces that refract impinging light giving angular dispersion of the radiation, provides greater radiation dispersion at shorter wavelengths than longer ones.
what determines the format of the final report?
type of project and by the end user of the data that one has generated
how long should samples that degrade over time be stores?
until their reevaluation is no longer possible
which test for urine shows ammonia indicated by production of deep blue color in test tube
urea nitrogen tube test
describe characteristics of urine under a microscope
urine contains solid materials with characteristic crystalline structures + epithelial cells
immunoassay
use of antibodies for the detection of chemical substances, based on competition between labeled drug and drug class in specimen for sites on antibody to drug class (competitive binding)
blood concentration
used for analysis of alcohol concentration on a living person
Developmental validation
used for new techniques should include: characterization of the locus, species specificity studies, sensitivity, stability and mixture studies # of samples from a pop must be obtained and analyzed in order for independence studies and allele frequency determinators to be made.
LC
used for polar or thermally labile compounds, easily changed to accommodate different analytes
UV-Vis Spectrophotometry
used in pharma and biomedical analysis for quantitation, characterization of drugs, diluents, metabolites, and other related substances. Technique works on the principle that substances absorb and emit ER at diff wavelengths. Abs and emit is recorded to provide a spectrum and used to deter the conc of a substance in a soltn
substrate molecules
used in some immunoassays to react with the molecular label and visualize the antibody-antigen pairing
What is the function of proteinase K in DNA extraction?
used to digest contaminating proteins
urine
useful for testing (human performance), may have higher concentrations than blood, drugs present for days, rapid screening
bile
useful in absence of urine, some drugs concentrate here for easier detection
confirmation test
uses both gas chromatography and mass spectrometry
Mass spec
uses different voltages to select out certain molecules
Trinder's test
uses trinder's reagent, contains ferric ions that creates color violet complex with salicylate ions
performance enhancing drug testing
usually split specimen - A for testing, B reserved for retests
can a lab refuse receipt of samples if the integrity of it is in question of there is inadequate, incomplete or inaccurate transmittal forms?
yes - they can also accept it but document the lack of security/inadequate documentation.
Blood Borne Pathogen Training
--all personnel should have blood borne pathogen training once a year if they are exposed to them --Hep B vaccinations must be made avail to an employee within 10 days of an assignment to a position involving blood borne pathogens --Treat all human bodily fluids should be treated as possibly ctg HIV, HBV and other blood borne pathogens --Gloves and handwashing --Equip and lab surfaces cleaned and disinfected. All waste labelled. Est areas of pre and post DNA amplification --All areas contaminated with biological waste must have fluorescent orange or orange red labels with the biohazard symbol
Preliminary (Presumptive) test advantages
--cheap, no expensive equip needed, minimal training, requires minimal sample
To prevent contamination there must be separate areas for:
--mixing of the test --control and reference substances with a carrier --storage of the test, control, and reference substance mixtures --areas for pre and post amplified DNA --extraction and amplification should be separate areas --examining reference samples and evidence should be done separately
screening uses
"initial tests/ medical tests", no false negatives, easy to perform, automated analyses, cut-off values, cross-reactivity
vitreous humor
(eye fluid) - used in various analyses (clinical chemistry, ethanol, drugs), more resistant to decomposition
Kinetic Interaction of Microparticles in Solution (KIMS)
(inversely proportional) microparticle labeled with several drug molecules, aggregates form when several antibodies bind to the drug, as aggregates form absorbance increases, with more drugs in the sample, they bind to antibody leaving the microparticle unbound. MORE drug = LESS aggregates
first order elimination
(most drugs) logarithmic relationship, concentration dependent and variable
What're two advantages to permanently fixed eyewashes?
- cannot be misplaced or removed - more likely to be periodically flushed
Items employee need to wear while conducting analysis are (clothing and protective gear):
- lab coat - items that will prevent contamination of test equipment, the test or control articles - items that will protect from potential dangers (biological, radiological, chemical
pH and toxicology
--absorption, distribution and excretion of drugs --extration and analysis for drugs --pH is temp dependant (20-30 C)
Probability should always = ____
1
How often does hair grow?
1 cm/month
Lividity appears _____ post death and fully develops ______. It helps with determining------
1 hour, 3-4 hours, if the body was moved
chain of custody prevents these 2 ways analytical results can be challenged:
1. showing the analytical procedure wasn't accurate; 2. raising doubt as to whether the sample really was the one that was originally obtained
how many hours post mortem does rigor mortis become fully present?
12
UV absorbs at what wavelength?
200-400nm
Nucleic Acids have UV-Vis absorption characteristics at which wavelength range?
240-280nm
How many calibrations are needed to construct a standard curve?
4-7
how many years must non-clinical study records be retained if an application is made for a research or marketing permit?
5 yrs
Rigor Mortis (rigidity) sets in about _____ hours after death and lasts _____ days.
6-12 hours, 2-3 days
Sample Log In to Lab
A chronological log of all samples should be maintained with the following info: date sample was collected, date sample was received, unique laboratory ID #, cross-reference to original field #, required analyses, signature or initials of person logging the info, comments on sample integrity ID #'s should be on sample ctgr not lids
What is a population database?
A collection of DNA profiles
Colored Crystals
A colored test reagent may result in the formation of
High Performance Liquid Chromatography (HPLC)
A form of chromatography in which a small sample is put into a column that can be manipulated with sophisticated solvent gradients to allow very refined separation and characterization; formerly called high-pressure liquid chromatography. Detectors: UV/Vis Advantages: most compounds absorb UV, sensitive to ng/mL Disadvantage: eluent absorb at optimum detection wavelength, need chromophore, poor sensitivity Fluorescence Advantage: improved sensitivity, more selective Disadvantage: few fluoresce naturally Electrochemical Advantage: sensitive to pg/mL, selective Disadvantage: high maintenance
Enzyme Multiplied Immunoassay Technique (EMIT)
A non-separation immunoassay based on an enzyme label Strength: Homogenous, long shelf life Weakness: matrix substances cross react, interfering substances
What are the ABO phenotypes?
A, B, AB, O
a protein produced by the body's B cells in response to the presence of a foreign body form the primary immune response, and function to resist disease by attaching themselves to the foreign antigen, and through a cascade of actions cause it to weaken or be destroyed
Antibody
How do the ABA Card P30 and Seratec PSA Semi-quaint work? Are they presumptive or confirmatory?
Antibody/antigen rxn between PSA & antibodies on test card. Sim to hematrace. Both work similarly; seratec has semi-quantitative internal control 4ng/mL * confirmatory *
bacterium or virus stimulates production of antibody
Antigen
Define bloodborne pathogen
Any disease causing microorganizm present in human blood.
Define Forensic Science
Application of science to law.
Cocaine, Crack Final Color, Dark Blue
Cobalt thiocyanate
Cobalt Thiocyanate
Cocaine, Crack, Procaine, Lidocaine
Cobalt Thiocyanate Test
Cocaine, Crack, caines Blue
What environment would not harm DNA? Heat, Humidity, UV light exposure, cold
Cold
Describe the proper way to collect and store the evidence below: Pooled blood stain
Collect with a sterile gauze after photographing, then freeze or refrigerate immediately
Fluorescent Polarization Immunoassay (FPIA)
Competitive, homogenous. Inverse relationship between fluorescent and Ag concentration. Limitation based on MW - good for low molecular weight antigens. Strength: homogenous, very stable, low limit of detection, less matrix effects in old blood and urine Weakness: Expensive, can give false positives, not adaptable
Closed enzyme-donor Immunoassay
Competitive, homogenous.Dimeric enzyme, subunit of B-galactosidase. Strength: linear over wide ranges, can measure second wavelength Weakness: new technology, troubleshooting difficult, affected by adulterants
When performing the sulfur test, what color precipitate should show a positive outcome?
Black or dark grey
When using postmortem toxicology testing what is the most important specimen to use?
Blood
What color does tetramethylbenzidine turn in a positive reaction?
Blue
include negative tests, reagent tests, and derivative tests.
Indirect Tests
How do you package evidence at a crime scene?
Individually, allow wet objects to dry
Disposal of Records
Information that can be used to ID a human must be destroyed, provisions may need to be made for the off-site storage of non-active and permanent records
solid phase extraction
It is a form of digital (step-wise) chromatography designed to extract, partition, and/or adsorb one or more components from a liquid phase (sample) onto stationary phase (sorbent or resin). Advantages: Selectivity, Reproducibility, Speed Disadvantages: Protein binding
What is the role if isoamyl alcohol in organic DNA extraction methods?
It reduces foaming caused by SDS
What salts are in the blood?
K, Cl and Na
Strong Acids (Ka and pKa value)
Ka >1, pKa < 0
What two scientists were known for devising experimentation for blood types?
Karl Landsteiner and Leon Lattes
Strong Bases (Kb and pKb value)
Kb > 1, pKb < 0
What is the major protein in hair?
Keratin
Which one of the following statements is true? A. Gunshot residue analysis of a firearm should take place after biological screening B. Tool mark analysis of a firearm should take place prior to biological screening C. Gloves worn to protect latent prints are typically made of latex D. Gloves worn to protect latent prints are typically made of cotton
D. gloves worn to protect latent prints are typically made of cotton
Surface tension [Increases/Decreases] the surface area of a drop of blood.
Decreases
can result in physical or chemical changes.
Degradation of drugs in the solid state (tablets)
What are 2 examples of physical properties?
Density, color, texture, odor
the drug-reagent derivative formed, becomes the compound to be tested.
Derivative Tests
Parts of a study protocol
Descriptive title (ID #, status (draft, pending), approval date) Study Objective (purpose of study-dose confirmation, dose determination ,concentration of by-product Identification of the Test and Control Articles (by name, chemical abstract # or code) Sponsor (indv, organization, or testing facility that supports the study and all their contact info) Testing Facilities (the person or laboratory that actually conducts the experiments) Others involved Justification for Study (info given to justify the study objectives ex. published work
Explain the difference between descriptive and inferential statistics.
Descriptive-using graphs and charts to organize the information Inferential-using information to make predictions
Entomology
Determine time of death using insects
Toxicology
Determine whether drugs are in a human body
include precipitation tests and color tests; the substance being tested is responsible for the crystals produced.
Direct Tests
- Emulsions may form trapping drug - Immiscible solvents are required - Time consuming - Glassware clean up required - Large volumes of solvents used - Operator dependent (some skill involved)
Disadvantages of Liquid-Liquid Extraction
- Lower sensitivity and resolution than gas chromatography and high performance liquid chromatography
Disadvantages of TLC
Name the formal procedure used by parties to a lawsuit to obtain information before trial.
Discovery
Specificity
Discriminating bt compounds of closely related structures that are likely to be present. This can be demonstrated by getting positve results in a sample that ctg the analyte and negative results in a sample that does not ctg the analyte
• Changes in ambient temperature and humidity • Concentration of the reagent or substance tested
Distortions of crystal form resulting in crystal di- or polymorphism, can be caused by:
What information should a company instrumental techniques of controlled substances?
Documentation Case #, Item #, and Initials.
the soluble drug-antibody complex is rendered insoluble by adding a second antibody (precipitating antibody targeted to the globulins (albumin) of the animal in which the first antibody was raised). The bound and free form can then be separated by centrifugation, the heavier double antibody complex forming a pellet that can be recovered and measured.
Double Antibody Technique
This is the most sensitive type of precipitin test that gives qualitative and quantitative measurements.
Double diffusion in one dimension
Alkaloidal base, primary, secondary and tertiary amines Final Color, Orange; red-orange; brown-orange
Dragendorffs reagent
Marijuana, hashish, THC Coffee (roasted) Patchouli oil Tea (leaves) Final Color, Violet Blue
Duquenois reagent
This is the technique that is used to separate charged proteins based on pH. The pH is low at the anode and high at the cathode.
Isoelectric Focusing
Decide which statistical test is associated with the info for each: used for a non-negative distribution and the critical value is right tailed
F-Distribution
T/F ID#s for samples can be placed on lids and other parts that can be separated from the sample
FALSE - place directly on sample
uses the difference in rotation of bound and free fluorophore-labeled drug.
FPIA - Fluorescent Polarization Immunoassay
Crime scene processing is the same as crime scene investigation.
False
T/F If a sample is degraded the chances of obtaining a mtDNA profile is less than obtaining a nuclear DNA profile.
False
T/F Microchemical confirmatory tests are not easily disrupted by debris and dirt, and are more sensitive than color tests
False
T/F Process both question samples and reference samples together.
False
T/F Random Match Probability is an exact number.
False
T/F The yield gel only measures total human DNA.
False
T/F You can analyze your data without an allelic ladder.
False
it is possible to individualize trace evidence.
False
T/F Amplified DNA does not need to be in separate area than DNA extraction and PCR set-up.
False (DAB standard 6)
T/F balances in a lab must receive monthly maintenance
False - yearly, plus zero check and standard mass reading with each batch of weighings.
T/F all areas contaminated with biological waste must have fluorescent yellow or pink labels with the skull and crossbones symbol
False -- fluorescent orange or orange red labels; biohazard symbol
T/F a chain of custody accounts for a sample from the moment it is checked in to the lab to the moment it is discared
False -- from the moment it is COLLECTED to the moment it is discarded/destroyed
This is an alternative to the Frye standard that is more flexible
Federal Rules of Evidence
Name a sample type in which Y-STRs would not be helpful.
Female victim was beaten by unknown individual, suspected to be her ex-boyfriend's new girlfriend.
What are natural fibers?
Fibers made from animals or plants
Friction ridges are located on our _______, palms and bottom of our feet.
Fingers
What are 2 examples of chemical properties?
Flammability, reactivity
What are the 4 search patterns for crime scenes?
Grid, quadrant or zone, spiral, line or strip
DNA analysis that is the same as VNTR but uses a reverse blot to visualize
HLA DQ Alpha
what type of chromatography typically determines chromatographic purity?
HPLC
Ehrlich's Reagent
Hallucinogens
Ehrlich's reagent
Hallucinogens Violet
what does Ehrlich's reagent test for and what color is considered positive?
Hallucinogens, *violet*
Standard Personal Protection Equipment includes what four elements?
Hand, foot, eye, mouth/nose protection
small non-immunogenic molecule attached to a larger immunogenic substance forming a new antigen that stimulates production of antibodies specific for the small molecule.
Hapten
The Chelex resin is selective for ____?
Heavy metals and cations
What is the name of the well known classification system for fingerprints still used today?
Henry System.
Derivatization for GC
Issue with GC is that it requires the analyte to be volatilized at an elevated temp. This process can breakdown chemicals susceptible to heat. A poor separation can occur if compounds are not easily vaporized. All these issues can be dealt with if a derivative of the drug of interest is produced. This can be done using a derivatizing agent.
FTIR sample prep
IR spectra can be obtained from vapors, liquids and solids. Most commonly used by toxicologist is the solid phase. Can either make then into a soltn or place them bt two disks
What do forensic scientists use physical evidence for?
Identification and comparison.
Describe the proper way to collect and store the evidence below: Semen stains
If they are dried you can either collect the entire fabric they are on or get a scraping. If they are wet you should allow it to air dry then place in a sealed paper bag. Do not subject the sample to heat or humidity.
techniques used to measure the presence and amounts of a chemical substance in body fluids such as blood, plasma, serum, vitreous humor and more commonly, urine.
Immunoassays
Acidity and basicity of solutions is conventionally measured using the pH scale. pH measurements are of importance in many areas of toxicology: • Absorption distribution and excretion of drugs • Extraction and analysis for drugs
Importance of the pH Scale
Pathology
Perform autopsies to determine death
What's the most common test for saliva, because of its sensitivity and specificity?
Phadebas
Every crime scene must be clearly documented by ____________ addition to notes and diagrams/sketches.
Photograph's
Involves no change in the covalent bonds of the drug but may involve changes in the crystal lattice structure.
Physical Decomposition
temperature, catalysts, pH and nature of solvent/s present.
Physicochemical factors that affect drug decomposition rates include,
presence of glandular trichomes containing orangey cannabis oil
Plant material that distinguishes the material as being derived from Cannabis sativa.
A type of ridge detail is a _______ print when ridge detail is indented into a substance.
Plastic
Besides establishing what is known as The Locard Principle, what else is Edmond Locard known for starting?
Police Laboratory
Real-Time PCR utilizes a modification of what method?
Polymerase Chain Reaction (PCR)
the pure drug can exist in more than one different crystalline form. Have different solubilities
Polymorphic Form of a drug
a substance that is known to react in the same way as a positive sample. For example, if testing a urine sample for the presence of amphetamine, a urine sample spiked with a standard solution of amphetamine will suffice as a positive control.
Positive Control
Applications of Forensic Toxicology include workplace drug testing, Investigation of contraband materials and ___________.
Postmortum
The short polynucleotide chains used in PCR to target regions of amplification are called ____.
Primers
EMIT pros & cons
Pros - less steps (homogeneous), easy to automate, long shelf life, easy to discriminate between large range of concentrations, reduced interference due to measurement over time Cons - potential matrix interference, potential false positives/ negatives, some substances can disrupt enzyme activity
FPIA pros & cons
Pros - less steps (homogeneous), long shelf life, fluorescein-labeled drug more stable than enzyme drug, low limits of detection, less interferences present in matrices compared to homo. enzyme assays Cons - more expensive than enzyme assay, potential false positives, not easily adaptable to high through-put analyzers
Type of DNA analysis where DNA is cut into fragments with a restriction enzyme that recognizes short DNA sequences.
RFLP
Temperature and Rxn Rate
Rate of a rxn is based on the temp. in K. The rate increases by 2-3 for each 10 C rise in temp.
What is the name for the information collected directly from the CCD camera?
Raw Data
a color change or precipitate formation is due to some change in the reagent caused by the test substance.
Reagent Tests
Computer Forensics
Recover data from digital media
For short term storage, liquid blood samples should be kept _____.
Refrigerated
What is a felony?
Major Crime
How should you collect fiber evidence?
Make tape lifts, bag evidence individually, removed fibers should be folded into a sheet of paper and stored in a paper bag
chemical, physical or microbial degradation.
Many drugs are not chemically stable and can be subject to
Duquenois Reagent
Marijuana, Hashish, THC, coffee (roasted), parchouli oil
compounds that contain basic nitrogen and readily precipitate from aqueous solution. These reagents are also reactive with compounds in which oxygen atoms have basic properties making them capable of forming oxonium salts.
Microcrystal tests are useful for the identification of
Direct and Indirect tests
Microcrystal tests for chemical identification purposes can be classified into
What's the hazard code symbol to detect dangers?
NFPA: National Fire Protection Association.
NIDA
National Institute on Drug Abuse, consensus document detailing conditions for urine drug testing
a drug free urine sample.
Negative Control
tests where the test is the failure of a specific action or reaction to occur.
Negative Tests
In the process of DNA electrophoresis, [negatively/positively] charged DNA moves [away/toward] the [anode/cathode].
Negatively charged DNA moves toward the anode
What is the difference between normal and cancer cells?
Normal cells replicate a certain number of times and don't continue to replicate. Cancer cells continuously replicate.
What type of DNA is found at the hair root?
Nuclear DNA
What three fibers dissolve in formic acid?
Nylon, silk, acetate
UV or Vis Components: Detectors
Often a photon transducer which has an active surface that absorbs radiation using a photocurrent or enhanced conductivity, the most common detector is a photomultiplier which is made of several electrodes called dynodes. The electrical potential bt dynodes increases causing acceleration of e- that are ejected from the photoemissive surface. When e- strikes the dynode, several electrons are ejected. Eventually there is a cascade of e-.
liquid-liquid extraction
Requires two immiscible liquids (aqueous & organic phase) use separatory funnel; lower density on the bottom; like dissolves like; organic non-polar compounds will dissolve in organic phase Disadvantages: Emulsions form Immiscible solvents required Time consuming Glassware cleanup Large volumes used Operator dependent
Possibility of Sample Contamination
One of the biggest concerns of any forensic laboratory and the point of attack for many defense attorneys
Marquis Reagent
Opium - purple Amphetamine, methamphetamine - Orange MDMA - blue to black
what protein in semen comes from the prostate and is especially important in semen samples of aspermic or oligospermic males?
P30
What provide a contained clean area to set up PCR reactions without expense of a biohazard hood?
PCR set up cabinets. they should have their own set of equipment- dedicated pipettes for use solely in these cabinets
DNA analysis that is used to analyze loci simultaneously
PM Polymarker
What organization prepares guidelines and proposals for minimum examination and identification of controlled substances.
SWG Drug
without allowing the column to dry out, the sample is applied and pulled through the cartridge. Then the column is air dried for approximately 30 seconds to ensure as much of the sample matrix as possible has been removed.
Sample Application (Load) (TLC)
This test is used on saliva samples, ABH antigens will neutralize corresponding antibodies.
Secretor Status
What are the 5 steps in crime scene investigation?
Secure crime scene Observe crime scene Document crime scene Search crime scene Collection & preservation of evidence
What can fracture patterns tell an investigator?
Sequence of events Direction of impact High or low velocity impact
How are MSDSs obtained?
Shipped w/ first order of chemical; can be requested from manufacturer or distributor @ time of purchase. companies have web-accessible collections; also available on some general websites
Components of a Final Report
Should include: title page, abstract or summary, highlights, reserach plan, procedural changes, analysis of data, storage of records, signatures and dates, corrections and additions to the final report
Decide which statistical test is associated with the info for each: This model describes the association between two variables and uses y=mx +b
Simple Linear Regression
Solid phase extraction cartridges consist of a polypropylene column containing bonded silica sorbent material held in place by an upper and lower frit.
Solid Phase Extraction
The antibody to the drug is bound to the wall of a plastic tube. After incubation, the drug-antigen is bound to the wall of the tube. The bound drug can then be analyzed by gamma counting the tube or by eluting the drug-antibody complex, resuspending it in solution, and measuring any optical changes.
Solid Phase Extraction Systems
Which search pattern is typically used underwater?
Spiral search
Injector for Gas Chromatography
Split mode--the septum purge remains open, this minimizes extraneous peaks result in a septum bleed Split-less mode--(split valve remains closed)use for low conc samples to get as much of the sample as possible onto the column
What is this fingerprint type?
Whorl, 2 Deltas.
Would you expect a blood stain resulting from a 90* angle to be WIDER or THINNER than that from a drop coming from a 50* angle?
Wider (thinner = more acute angle)
What well known incident example ended the use of the Bertillon measurement system and convinced a police agency that fingerprints were a superior method of personal identification.
Will & William West
This is mostly used in rape and paternity cases bc it uses the Y chromosome found in males
Y-STR
Can scientists determine body area that hair came from comparison?
Yes
which case does not generally require an autopsy?
a 78 year old heart attack victim who died at the hospital
What pattern does a fire burn?
a V
Inspection
all items used in an analysis must be inspected, equip qualification is a type of inspection, equip and consumables must be examined after purchase, identities of items should be verified and documented, other forms included would be performance qualification, calibration, and tuning procedures, inspect glassware for cleanliness and cracks
solutions of drugs or chemicals of known concentration
Standard Solution
Decide which statistical test is associated with the info for each: This test is used with normal distribution, 2 tails extend to infinity, sample size should be greater than 30
Student's t-distribution
GABA receptor
allows chloride channels to pass- hyperpolarizes cell - decreases likelihood of an action potential
Which of the following are examples of ionizing radiation? IR, alpha, x-rays, beta, Vis, sound, radio, gamma
alpha, x-rays, beta, gamma
Skeletonized blood samples, dilution by snow or rain and clots of blood are examples of _______blood stains.
altered
half-life
amount of time it takes a drug to reach half its concentration
Schedule IIN
amphetamine, methamphetamine, methylphenidate, amobarbital, glutethimide, phenobarbital, PCP
Hairs that have naturally fallen from the head are what category? Anagen, Telogen, Catagen
Telogen
The rate of a chemical reaction is related to the absolute temperature in Kelvin (Arrhenius). Rates increase by a factor of 2-3 for every 10°C rise in temperature. The Arrhenius equation is applicable to solution reactions providing that the temperature range is between 30-50°C.
Temperature
the basicity of the base that accepts the proton. The basicity of any base depends on the acidity of the acid that donates the proton.
The acidity of any acid depends on
Factors that affect Quality Assurance and Quality Control
Turn-around time, quality and price Depends on the industry as to which is more important For food industry it's turn around time ex. produce industry works on positive release basis (results prior to distribution) There are situations when both are needed and this is a challenge ex. gov. commissioned food safety and quality monitoring programs Price is the rate offered to the customer. Loss leaders are when a item or service is sold below cost to attract customers
By using profiler and cofiler kits, how many amplifications does it take to obtain information on all the CODIS loci (currently 13)?
Two
TLC is one of the most widely used techniques for the separation and identification of drugs and can be applied, to pure drug, drug extracts and extracts of body fluids and tissues. This technique maintained popularity due to its simplicity, reliability, cost effectiveness and selectivity.
Thin Layer Chromatography (TLC)
What is the LOQ?
This is the level of quantitation, the lowest level at which the instrument can detect the analyte
IR Vibrational Modes
Three types of absorption occur: 1. Fundamental--primary abs bands for each mode of abs 2. Overtone--occur at wavelengths that are the sum or diff bt two or more fundamental bands 3. Combination
where are SOPs for analysis of FDA-approved drugs found?
USP/NF: united states pharmacopoeia/national formulary and the homeopathic pharmacopeia
T/F 55% of blood contains no cells.
True
T/F Random Match Probablity is calculated from the frequency of DNA markers in a database.
True
What is the fundamental mechanism considered necessary to maintain the BBP standard and are the primary mechanisms for infection control?
Universal Precautions
Friction ridges develop during fetal growth and remain unchanged throughout life until after death.
True.
Lab Receipt of Sample
Upon receipt of a lab sample the laboratory should: verify the integrity and condition of the sample ctgrs, verify documentation (ID #s match, all samples present), verify storage temp
What type of specimen is most frequently used in pre employment screening?
Urine
T/F: SpermPaints fluorescent dye w/ antibodies bind to sperm head and tail antigens. Selective only for sperm and won't stain anything else.
True. use in conjuction w/ laser micro-dissection microscope.
T/F One a person with appropriate quals has been hired, the lab must provide them with training on specific tasks he/she will perform
True.Furthermore - employees are expeted to demonstrate competency for each analytical procedure they perform
What types of swabs are most common in sexual assault kits?
Vaginal, oral, anal swabs
factors effecting Vd
age, disease, gender, weight
Methylating agents for GC (used for derivatizing drugs)
agent used to derive barbituates and some carboxylic acids. Methylation can be achieved by using HCl in methanol.
What does dragendorffs reagent test for; what is the final color?
alkaloidal base, primary, secondary & tertiary amines; *orange*, *red-orange*, *brown-orange precipitate*
Checking for homogeneity of soltns
an aliquot should be taken from the top, middle, and bottom of the sample ctgr. each sample is analyzed X3, conc. should fall into previously deter. acceptable range for sample to be considered homogeneous
sensitivity
assay characteristic that denotes the assay's ability to DETECT an analyte in the sample
specificity
assay characteristic that denotes the assay's ability to correctly IDENTIFY an analyte in the sample
Examples of drugs that are bases
atropine, codeine, epinephrine, gentamicin, streptomycin, cocaine, BZE
Common Descriptive Statistics
average (or mean), median, mode (appears most often), confidence interval (how confident one is that results are accurate), measures of precision (range, standard deviation (scatter of values around the avg value) , variance (best measure of precision), relative std deviation or coefficient of variation, graphical representations, calibration curves
rectal
avoids liver/first pass effect, good for non-compliant situations, stigma, inconvenient
what is detrimental to the survival of DNA?
bacteria, heat, moisture
writing instruments include
ball point pens, spray cans, paint brushes
which is not a type of glass discussed in class?
borosilicate
in forensic biology, confirmatory tests
can determine a sample to the exclusion of all other possibilities
instead of separate storage for each lab, what's a more efficient and cost effective model?
central stockroom
colorimetry
chemical reactions resulting in visible color changes
Bullets give (class, individual) characteristics
class
when a spot test of cobalt thiocyanate turns blue, what compound is suspected, and what should be done next?
cocaine or crack cocaine. achieve confirmation by IR or GCMS.
What does cobalt thiocyanate test for & what is the final color?
cocaine, crack, 'caines; *blue*
oral
convenient, faces absorption challenges, subject to first pass effect, slow onset
Computer Tracking--Analytical Information Management System (AIMS)
converts data from standard format, search data archives and serves as a central repository for all instrumental data
Schedule V
cough preparations with </= 200 mg codeine/100 ml or g, ezogabine
repeating arrangement of individual molecules (crystal lattice) and a characteristically narrow melting point range.
crystalline drug
UV or Vis Components: Sample Cells
cuvettes must be transparent to incident radiation. usually made of quartz or fused silica bc they are both transparent to UV and visible range wavelengths, most common cell length is 1 cm, the same type of cuvettes should be used for measurement of reference and test samples, also don't the outer surface with fingers to avoid oils
immunoassay
depends on drug-antibody reactions
alcohol is an example of a
depressant
Medical Review Officer (MRO)
licensed physician who has knowledge of substance absue disorders and is trained to evalutate and interpret results, liason between labs/employment/donors
forensic urine drug testing
determines the absence or presence of drugs and their metabolites in urine to demonstrate prior use or abuse
active metabolite
like the parent drug, still has an effect on the CNS
when might employee illness contaminate or bring into question integrity of analysis?
drug studies .the employee must be removed from the study until the condition has been corrected.
acids or bases
drugs are chemicals that are classified as either _____ or _______
First pass effect
drugs are destroyed in the stomach first, this decreases the concentration of the drug before metabolism even begins, and liver can metabolize before drugs reach systemic circulation
what bacteria in feces are strong presumptive evidence?
enterococci
What is another name for RBC's?
erythrocytes
whats the most frequently used controlled substance?
ethanol
which is the most abused drug on the world
ethanol
monoclonal
every antibody has identical binding properties
Chemical reactions that give off heat are called
exothermic reactions
Extraction Efficiency
extraction concentration of sample divided by the concentration of the spiked analyte Ex. urine extracted 400ng/ml and urine spiked with 500ng/ml, so 400/500 = 80%
percentage of alcohol content
factor effecting absorption and distribution
The lowest temperature at which there is enough vapor to be ignited is called the
flash point
the most common instrument used to give an indication of the type of accelerant used to start a fire is
gas chromatography
confirmation test
gas chromatography is completed first; then each separated component enters the mass spectrometer where the sample is broken into fragments for identification
in order to have a flaming combustion, both the fuel and the oxygen must be in the
gaseous state
what contain brown-orange oil that distinguish plant matter as being derived from cannabis sativa?
glandular trichomes/hairs
name 3 items of personal protective equipment (PPE)
gloves, lab coat, face shield, gown, respirator mask, shoe covers, etc.
tests for identifying vaginal secretions screen for what?
glycogen
Study Protocol: Experimental Design
goal is to obtain data in a statistically relevant manner, the analyses must be shown to be reproducible, large sample size, variables controlled, measurement more useful than observation, how subjects are selected must be discussed, ensure internal validity of the data by reducing bias by using the double-blind technique and cross-over experiments.
Which of the following is a search pattern?
grid
a blood spatter pattern consisting of a large number of very fine, mist-like droplets indicates that the blood probably came from a
gunshot wound
angel dust or PCP is what kind of drug
hallucinogen
Laboratory Notebooks
hardbound logs that are numbered, information is recorded by hand, notebook pages must be consecutively numbered before using the notebook, all data must be entered in indelible (permanent) ink, date, time, signature/intitials should be written after each major step, printouts from instruments or photos of gel-electrophoresis can be included
thin layer chromatography
has a solid and a liquid phase; separates molecules by density
absorption
has occurred when alcohol appears in the blood stream
the method for separating and concentrating ingnitable liquids that uses a syringe to collect samples is the
heated headspace method
The total volume of the RBC's is called the _____.
hemacrit
what drug gives a characteristic purple color with marquis spot test reagent?
heroin
Schedule I
heroin, LSD, marijuana, peyote, methaqualone, MDMA/MDA/MDE
Radioimmunoassay (RIA)
heterogeneous (inversely proportional) - uses iodide to label drug, uses radioactivity instead of absorbance to determine presence of drug, MORE free drug = LESS radioactivity
Enzyme-Linked Immunosorbent Assay (ELISA)
heterogeneous assay (inversely proportional)- antibody is immobilized (coated wells), sample diluted in buffer, enzyme-labeled drug is added, incubate, wash, add substrate to change color, incubate, add stop reagent, read absorbance. MORE color = less binding of drug sample, more binding of labeled drug, therefore LESS color = more drug binding, less labeled drug
A gunshot injury involves ________ velocity blood spatter.
high
what type of radioactive energy emitters may require lead-lined drawers and bricks to place in front of an experiment?
higher energy gamma-emitters
______ of a preparation of suspension or solution of reference standards should be checked before using it in quantitative analysis
homogeneity
enzyme-linked immunosorbent assay (ELISA)
homogenous assays Strength: good sensitivity, less prone to matrix effects Weakness: cost per sample high, sodium azide blocks horseradish peroxidase action, trouble shooting difficult
handwriting comparisons can be made using
how letters are made, where pressure is applied, and connections between letters
blood spatter patterns can indicate
how many sources of blood there were, what type of weapon was used, the number of times a weapon was used
Schedule II
hydromorphone, methadone, meperidine, oxycodone, morphine, opium, codeine, hydrocodone
In relation to confirmation of blood, describe how spectroscopy works
identifies between Hgb and Hgb derivatives like methemaglobin, carboxyhemoglobin, sulfhemoglobin and oxyhemoglobin. all derivatives exhibit a strong absorbance at 400-425 nm (soret band)
Point of care testing (POCT)
immediate results, beware of limitations, send out for confirmation
How to avoid false positivies and negatives
include positive and negative controls in the testing regimen, these controls should be prepared by another analyst to prevent contamination
excitatory aa neurotransmitters
increase excitability of the target neuron, activate receptors that depolarize the post-synaptic membrane
describe the way the urea litmus paper test works
indirectly indicates presence of urea by reacting to urease, generating ammonia from urea. Ammonia indicated using litmus paper (basic - pH).
which is not one of the categories of patters we discussed in chapter 4
individualizing, reconstruction, sequencing. and these are all patters for analysis
a substrate control sample helps to detect
interference resulting in false negatives/positives
Pharmacodynamics of cocaine
interferes with reabsorption process of neurotransmitters, prevents reuptake of dopamine by blocking transporter, results in increased extracellular dopamine concentrations, produces euphoric rush, blocks sodium channels (decreases action potentials)
Fourier Transform Infrared Analysis (FTIR)
interferometer instead of monochromator, sample absorbs different wavelengths, interferogram will display the wavelengths that weren't absorbed Advantages: Rapid data collection, scan sample 10 times a second, all frequencies detected, increased sensitivity, increased resolution, enhanced data manipulation
how would you interpret the ABA test on the page?
invalid
you are using phenolphthalein to test a reddish brown stain for the presence of blood. When you put a drop of phenolphthalein on the swab with your sample, it turns pink immediately. This test is:
invalid because it turned pink
this volatile substance may require a fume hood having leaded glass and a HEPA filter
iodides
chemical methods for the development of latent fingerprints include
iodine fuming
Why is ALS useful in detecting semen stains?
it has fluorescing properties due to bacteria and flavins present. Visualized using 400-500nm wavelengths and using orange goggles in a dark room
what can affect how you prepare to conduct a crime scene investigation?
location, weather, time of day
when collecting document evidence, it is important to carefully protect the document to avoid
loss of possible DNA, loss of possible indentations, loss of possible latent print evidence
What is another name WBC's?
luekocytes
name 3 presumptive tests for blood
luminol, kastle-meyer (phenolphthalein), Leucomalachite Green, o-tolidine, benzidine, tetra-methyl benzidine, hemastix
TLC: Mobile Phase
make-up of this phase is dependent on the class/classes of drugs to be separated and the type of stationary phase chosen. Ion pairing agents can be added for separation of drugs that are strong anions or cations. The solvent should be pure and non-reactive with the drug substances being analyzed.
the polymerase chain reaction (PCR) is a process which
makes many copies of a specific piece of DNA
shed head hair has
mtDNA only
When the mean=median=mode is a _____ distribution but when the mean, median and mode are NOT equal then it's a _____ distribution
normal, skewed
Routes of Administration
oral, inhalation, intravenous, intramuscular, rectal, sublingual, intrathecal, transdermal, intraocular, intranasal, subcutaneous, topical
individualizing friction ridge detail also exists on your
palms, fingers, and toes
regarding storage,what media has a longer life expectancy - electronic storage or paper/microfilm?
paper and microfilm(50-200 years) - vs magnetic tape and CDrom, 5-20 years and2-100 years respectively
what should be used for examining pipes for traces of cannibis residue?
petroleum ether
genetic differences among people that enable them to be distinguished are called genetic ___?
polymorphisms
Results in a greater error than the sum of all errors by laboratory analysis alone
poor sampling protocols
Schedule III/IIIN definition
potential for abuse lower than I or II; may lead to moderate/low physical dependence or high psychological dependence
what makes separate stockrooms/storerooms mandatory?
potential toxicity and flammability of chemicals.
Why are separate areas set aside for mixing of tests, control & reference substances,storage of test control and reference mixtures?
prevention of contamination and human error
confirmation uses
primarily for forensic purposes, increased specificity, increased sensitivity, no false positives, utilizes second analytical procedure
liver
primary site of drug metabolism, ratio between blood and liver conc. may help determinations (manner/cause?), potential for higher conc. of both parent and metabolite
Analysis
prior to analysis the sample should be stored in a secure location with an unbroken seal, dilutions, measurement of pH and weight, duration time for processes such as extractions, amplifications, prep vol of reagents and stds., each step must be documented, for confirmation purposes another analyst should be there to observe OR another confirmatory test can be performed, all evidence should be documented with a sample ID #, date of analysis, ID of instruments used, approved methods number
intentional adulteration
process of adding an impurity or inferior substance ex. food industry adding fillers, selling frozen fish as "fresh"
polyclonal antiserum
produced by animals, ctgs many different antibodies with diff affinities for the antigen or compound of interest
Schedule III
products with </= 90 mg codeine/dose unit, buprenorphine
Sample stability/storage
protect samples from moisture, heat and light. Tablet/Powder storage--kept in a dry place, protected from light, and at low stable temps Liquid sample storage--store at an appropriate temp, protect from light and air Chemical degradation can occur and result in loss of pure drug by breakdown to another compound ex. cocaine to the metabolite benzoylecgonine Physical Degradation can also occur which can result in: changes in the crystal form, aggregation of drug particles in a suspension, loss of volatile chemical compounds to the atmosphere
regarding biological laboratories, what're 2 purposes of HEPA filters in safety cabinets?
protecting lab personnel from possible infections; protection biological material (cell structures) from infections carried by personnel
Reactions in which an acid reacts with a basic solvent or in which a base reacts with an acidic solvent are known as
protolysis or protolytic reactions
Computer Tracking--chemical information management system (CIMS)
provides the ability to store and work with chemical structures and rxns
drug dependency is subdivided into two categories
psychological and physiological
the process by which solid materials are decomposed by heat, forming smaller molecules that can support flaming combustion is called
pyrolysis
"Internal validation studies must be sufficient to show the ________ of techniques as practiced by that lab"
reliability
drug accumulation
repeated use of a drug will lead in many cases to accumulation of the drug leading to faster absorption than elimination, steady state reached about 5 half-lives of the drug (chronic v. acute administration)
Validation of Analytical Procedures: Quality Control
replicate control samples are spread evenly throughout each batch so that a control sample is injected at least every 10 samples and this is done at least 3 times.
Sensitivity
reported by reference to signal-to-noise ratios. Only applicable to analytical procedures that have baseline noise. Deter by comparing measured samples with known low conc of analyte to those of blank samples and establishing a minimum conc at which the analyte can be reliably detected. Ratio of 3:1 or 2:1 is considered acceptable
heterogeneous assay
requires bound and free antigen to be separated before labeled antigen is measured
cross reactivity
response to a substance other than the analyte of interest, potential for false positives
Storage of test articles
samples should always be stored to minimize degradation, loss, and contamination, they also should be secured to prevent tampering. The storage document must be linked to the sample analysis document. Disposal of samples must be documented, for DNA samples a portion should be retained
according to lecture, what is the first thing a first responder should do at a major crime scene?
search for and detain the suspect
Advantages of solid phase extraction compared to liquid-liquid phase extractions
selectivity, reproducibility, speed-automation
name advantages and disadvantages of color tests for drugs
sensitive, fairly quick; non-specific, destructive and may be affected by conditions of the test
Diff. bt sensitivity and specificity.
sensitivity = being able to deter low levels of the analyte specificity = being able to distinguish diff. compounds
What should you consider when validating a process?
sensitivity, specificity, accuracy, precision, quality control, linearity, extraction recovery
gas chromatography
separates out molecules that move up a thin coated plate; has a moving liquid and moving gas phase; the distance each component travels is based on the characteristics of that substance; compared to known samples
thin layer chromatography
separates out molecules that move up a thin coated plate; has both a solid and liquid phase; the distance each component travels is based on the characteristics of that substance; compared to known samples
solid-phase extraction
separation of the analyte from the matrix using a column, separates based on physiochemical differences
postmortem blood
should be collected from a variety of body sites
you have the same mitochondrial DNA as your
sibling
sensitivity is generally reported by reference to what ratio?
signal-to-noise
a person's ____ is usually more distinctive than their normal handwriting
signature
what might a "wet chemistry" lab require vs. instrumentation lab?
sinks, bench tops, electrical outlets, hoods /// more electrical power, fewer sinks and hoods; access to 4 sides of instruments
What two factors affect retention time?
size and mass of molecule
4 crystalloptic parameters that can be used to ID or distinguish drug microcrystals
size, form, color and aggregation characteristics
identification tests should be able to discriminate between compounds of closely related structures that are likely to be present. this refers to a tests _________
specificity
Descriptive Considerations
statistic techniques that collect, organize, and analyze the data without drawing any conclusion or inference, used by labs performing routine analyses (ex. level pesticides in soil, creatine in blood, controlled substance in sample) Assumptions made include--data having a bell-shaped curve (Gaussian distribution), errors that random and independent of outside influences
Preparing for External Inspection
steps to prepare include: laboratory clean-up, review each facet of the lab operation for compliance, organize each operation for easy review by the auditor
Performance enhancing drug testing - drugs to look for
steroids, stimulants, opiates, diuretics and masking agents, opioids, peptide hormones, marijuana and synthetic cannabinoids, gene doping, others
when a fracture line in glass hits a pre-existing fracture line it....
stops
What can anthropologists gain from each of the bones below: teeth pelvis long bones skull
teeth- ID of person, age pelvis- sex long bones-height skull- sex, race
Modification of an SOP
temporary adjustments are needed for an SOP due to: --A specific grade of solvent or reagent is currently unavailable --Impurities if a sample may require an additional extraction step --The analysis includes additional analytes When SOP's are changed a single line should be drawn through the changes and initials (of the modifier) must be placed next to line
For linear relationships (bt conc and response)
test results should be eval by appropriate statistical methods. Ex. by calc of least squares regression line, min of 5 conc is recco to establish linearity
absorption
the act or process of absorbing the drug from the site of administration to the bloodstream
pharmacokinetics
the action of the BODY on the DRUG - absorption/distribution/metabolism/elimination
pharmacodynamics
the action of the DRUG on the BODY, medical use
metabolism
the body's ability to break down chemicals
metabolism
the body's ability to breakdown chemicals
parent drug
the compound that is taken into the body
antigenic determinant
the conjugated drug - the portion of an immunogenic molecule that binds to an antibody-binding site
class characteristics of a firearm detectable from a fired bullet include
the direction of the twist of the barrel of the gun
What does it mean in an arson investigation if the hinges have soot on them?
the door was open when the fire started
haptens
the drug - small, nonimmunogenic molecule that is attached to a larger immunogenic substance, forming a new antigen that stimulates production of antibodies specific for the small molecule
pesticide tolerance levels
the enforceable limit of pesticide residue that is allowed on food items
what document summarizes the work that has been done and provides to interested parties information that might not otherwise be available, and documents a lab has accomlished tasks for which it was funded?
the final report
the caliber of a weapon is
the inside diameter of a gun barrel
toolmark examinations may provide the following information
the kind of tool used to make the mark
Prior to applying an immunoassay technique in the lab
the lab should verify and establish a limit of detection and linear range for the method.
.04%
the legal blood alcohol concentration level for commercial drivers is _____
.08%
the legal blood alcohol concentration level for noncommercial drivers is ____
breath testing
the most widespread method for testing intoxication; it requires no less than 1.1 to 1.5 liters of breath to ensure that "deep-lung" breath is measured
the four components of a firearm cartridge are
the primer, the propellant, the cartridge, and the barrel
Due diligence
the process by which data is generated to demonstrate compliance with statutory limits is commonly referred to as this
why have so many routes of administration?
the route depends on many factors: disease, direct to site of action, time of onset, properties of the drug, convenience, safety route also depends on: immediate effects, delayed absorption, first pass effect, enterohepatic circulation, and distribution
pharmacology
the science of substances used to prevent, diagnose and treat diseases
Equipment File
there should be an inventory of all equipment in the laboratory, they should all have a unique ID #, and have a file The equipment file should include: --equipment qualification --calibrations --preventive maintenance --performance qualification testing
which of the following is not a hallucinogenic drug: marijuana, PCP, or LSD
they all are
Extraction Recovery
this is deter by comparison of the peak area observed for non-extracted std soltn in an appropriate solvent injected into the chromatographic system, to the peak area observed with a soltn prepared at the same conc. in the appropriate matrix, extracted and injected into the chromatographic system. This procedure is performed at least 3 diff concentrations with several tests at each conc.
Give an example of a cause of death.
this is the initiating reason that the person eventually died.
name the 9 components of the final report
title, abstract, highlights, research plan, procedural changes, analysis of data, storage of records, signatures and dates, corections and additions.
6 elements of an 'in house' SOP
title, author, issue date, ID number, page x of y, approval authority
What is the main function of RBC's?
to transport oxygen
confirmation
two individual tests with two separate techniques to identify a drug (screening + confirmation), mass spec especially useful, quantitation may also be used
Issues with Preliminary (presumptive) tests
usually very sensitive so false positives can occur. This is when a test identifies a drug as present and it actually isn't. Color tests (presumptive tests) are not specific but rather ID drug classes.
What causes false positives for an acid phosphatase presumptive test?
vaginal fluids, male urine, hygene products, plant matter, spermicides
Establishing Precision
validation of tests for assay and for quantitative determination of impurities, std deviation, coefficient of variation, confidence interval replicate analysis of control samples is generally performed on the same day and over at least 2 weeks expressed as the relative std deviation of the intraday and interday replicate analysis for each control sample.
how would one test drug paraphernalia for possible cocaines left behind?
wash out with methanol and residue should be subjected to TLC, UV or GCMS analysis for traces
Elutropic Series
way to deter solvent strength-- hexane--least polar.........methanol....water....--most polar
What does deci mean?
1/10
What type of fibers contain proteins?
Animal fibers
What is the only fabric to dissolve in acetone?
Acetate
If you are a secretor, what antigens would you secrete if you were type O? A. A, B. and H antigens B. O and H antigens C. H antigens only D. O antigens only
C. H antigens only
A grand jury is made up of _____.
Citizens
What anticoagulant compound was found in blood samples in the OJ simpson case?
EDTA
Name 2 bloodborne bathogens
Hepatitis B virus (HIB), HIV
A preliminary scene survey is commonly reffered to as a "__________".
Walk through
Single Beam Spectrophotometers
a single light source pass through a test sample to a single detector and offers advantages of greater energy throughput, superior signal to noise ratios and less cluttered sample compartments
what happened to the initial suspect in the Yates murder case?
he began a scholarship fund after winning a slander lawsuit
Chemical Inventory should include
name of chemical, approx qty, storage location, date acquisition, manufacturer, qty used, exp date, hazards, CAS #
manners of death
natural, accidental, homicide, suicide, undetermined
This is the protein in semen
p30
the buildup of lactic acid that causes muscle stiffness post mortem is called
rigor mortis
positive control
substance that is known to react in the same way as the positive sample ex. if testing urine for amphetamines, using a positive control that is spiked with amphetamines in the std, for the negative control there should be NO amphetamines (drug free)
T/F The target amount of extracted DNA necessary for PCR is in the region of 0.5-2.0ug.
False, it's ng
Extraction Sequence-(solid phase extraction)
--Wetting (conditioning)--sorbent must be wetted to produce an enviro suitable for the reproducible retention of the isolate.Done by passing solvents like methanol, acetonitrile and isopropanol through the column. --Changing of pH Enviro--buffer is passed through the extraction cartidge to introduce a pH enviro conductive to the retention of th analyte of interest onto the column. Ex. if a cation exchange column is used for the extraction of basic drugs, an acidic enviro will cause the drug to become positively charged and therefore retained by the column --Sample Application (LOAD)--don't allow the column to dry out, the sample is pulled through the cartridge, the column is then air dried for about 30 sec to be sure the sample matrix has made it's way out --Rinse (wash)--endogenous compounds present in the sample matrix, as well as analytes we have no interest in can be removed by washing the extraction cartridge with a suitable solvent. Any combo of aq/organic solvents can be used --Elution--analyte of interest is eluted from the column and collected into a tube using a suitable solvent. The strength of the eluting solvent can be adjusted to optimize the elution of the analyte of interest while at the same time minimizing the amount of endogeneous material removed.
Laboratory Ventilation
--all lab areas should have ventilation that replaces the air in the room with air that is non-laboratory air --4-12 air changes per hour --air intake should NOT be near the air exhaust --qual and quant of air ventilation should be analyzed every 3mo. --volatile chemical use requires fume hoods that are vented to outside --there should be 2.5 ft per every 2 people of fume hood --perchloric acid requires a special fume hood --hoods should have a continuous flow-monitoring device that is checked prior to and during use. --when dealing with DNA Class II biohazard cabinets should be used (these should be located away from walkways to minimize external drafts)
Water Usage in Laboratories
--each lab must have one sink for washing hands, this can also be used for washing glassware, etc --each lab should have an eyewash station --each lab should have a safety shower with a drain under the shower to aid in clean-up --plumbing should be made of acid-resistant materials --DNA analysis requires pure water --for post-amplified DNA, deionized water should be used
Bio-Materials Lab
--lab should have restricted access--this minimizes the # of people exposed --equipped with biological safety cabinets that have HEPA filters to protect lab personnel and protect cell cultures --changing into other clothing may be needed if working with dangerous biological material
Animal Care Facilities
--one species per room (aquatic animals exempt) --seperate rooms should be used for studies involving biohazards, volatiles, aerosols, radioactive or infectious. --animals should not housed within individual laboratories --sick animals should be isolated --enviro of the room can be altered according to the study protocol. Includes temp, humidity, lighting --storage areas for feed and bedding are required --access should be limited to personnel
Examples of General Standard Operating Procedures (SOP's)
--prep of bleach soltn --Prep of soltns --Deter sig figs --Receipt and Inspection of Sample/test article --use and operation of balances, pH meters, fume hood, measuring pipettes --preventive maintenance --limitations of method
Radioactive Materials lab
--special room design --may require Pb lined drawers and Pb bricks to place in front of experiment --Geiger counters should be on hand --Fume hood with Pb glass and HEPA filter needed --check for radioactivity on surfaces regularly
Study Protocol-Experimental Plan
--this includes: the #, weight range, sex, source of supply, species, strain, sub-strain and age of all animals used --IACUC approval-use of animals must be reviewed and approved --info presented to the IACUC would include: conditions, descript of animal cages, feeding, provisions, treatments and experimental use, disposal of study animals, disposal of animal by-products, procedures for adverse events. --use of human subjects requires Institutional Review Board (IRB) to approve --#, sex, age, and criteria for selection of human subjects. use of fetuses, children, pregnant women, and prisoners is restricted --An informed consent form must be read, signed and dated by all human participants. Must be written so all participants understand it. --Procedure for ID to the system, using specific criteria to select the test system
what're the specific stains used in christmas tree stain?
. Nuclear fast red for Heads (DNA), counter stain of PIC for tails (blue-gray)
Validation of a new DNA STR system
...
What is the range for pH?
0-14
What're the NFPA degrees of danger?
0-4,4 being most severe.
Chain of Custody Documents
1. a transmittal document to describe field activities and receipt of the samples 2. a document that records storage and disposal of activities 3. a document that follows the sample through its analysis
3 steps to metabolism
1) absorption 2) distribution 3) elimination
List (5) areas within a forensic crime laboratory.
1. Chemistry 2. Questioned Documents 3. Toxicology 4. Firearms 5. Biology 6. Latent Prints
Give 2 examples of the materials used for collecting the three-dimensional footwear or tire impressions.
1. Dental Stone 2. Microsil
What is an example of a class characteristic in the examination of footwear impression?
1. Design 2. Shape 3. Size
what are the 4 steps of instrument qualification?
1. Design qualification (DQ) 2. installation qualification (IQ) 3. operational qualification (OQ) 4. performance qualification (PQ)
List a crime scene search method.
1. Grid 2. Spiral 3. Zone 4. Line
List 2 responsibilities of a first responder at a crime scene.
1. Human Life 2. Secure the Scene
Poor Quality Management can result in or be the cause of...
1. Lacking attention to detail 2. Understaffed 3. poor workload management 4. non-validated tests 5. quality control and assessment 6. poor sample control 7. poor results verification
List 2 plants that are considered controlled substances that require a botanical examination as part of the identification process?
1. Marijuana (weed) 2. Peyote (cactus plant)
List 2 factors that can affect handwriting.
1. Old Age 2. Parkinson Disease 3. Arthritus
What are the 2 classes of writing standards utilized for comparison purposes?
1. Requested 2. Non Requested
List 2 types of firearms that may be submitted to the laboratory for examination.
1. Revolver 2. Pistol
What are the 2 basic types of crime scene sketches?
1. Rough 2. Finished
List 2 examples of surfaces that create three - dimensional footwear or tire impressions.
1. Sand 2. Snow 3. Soil
A Forensic Scientist working with Questioned Documents has many duties provide 2 duties that were discussed in class.
1. The pattern in the strokes of the lines and the amount of pressure of the pen touching the paper. 2. Examine for False document and insurance fraud.
List (5) examples of the use of the forensic sciences in the Sutherland case.
1. Tires 2. Hair 3. Fibers 4. Foot Impressions 5. Biology
What are the 5 manners of death?
1. accidental 2. natural 3. homicide 4. suicide 5. undertermined
Type of Detectors for GC
1. electron capture detectors---selective to electronegative compounds, especially chlorinated,fluorinated, or brominated molecules. Works via Nickel-63 emitting e- (beta particles) that collide with and ionize the gas particles. 2. flame ionization detectors--most sensitive gas chromatographic detector for hydrocarbons such as butane or hexane, based on the detection of ions formed during combustion of organic compounds in a hydrogen flame. 3. thermal conductivity detectors--The TCD compares the thermal conductivities of two gas flows—pure carrier gas (also called the reference gas) and carrier gas plus sample components (also called column effluent). 4. mass spectrometer--In order to measure the characteristics of individual molecules, a mass spectrometer converts them to ions so that they can be moved about and manipulated by external electric and magnetic fields.
Causes of non-linear results from UV-Vis
1. electrostatic interactions bt molecules in close proximity 2. scattering of light due to particles in the sample 3. fluorescence or phosphorescence of the sample 4. changes in the refractive index at high drug conc 5. non-monochromatic radiation 6. stray light
name 3 variables in human subjects that make clinical studies difficult and prone to uncertainty?
1. genetics. 2. lifestyle/exercise 3. environment 4. other medications 5. extent to which individual follows protocol
Methods to avoid obsolescence of data
1. keep original software, operating system, and all associated hardware 2. program all future computers to emulate the original software and operating system 3. Copy the record from the old format to the new one-migration
when must statistics be considered in a research project? (2 phases)
1. prior to conducting the study -- statistical approach that will give the most meaningful results must be determined; 2. once data is acquired, descriptive and inferential statistics are used to interpret data
Two important aspects of sampling
1. sample should representative of the entire amount 2. is there heterogeneity of the analytes in the batch
What are the 3 ways a criminal action can be initiated?
1. written complaint by law enforcement officer 2. Grand jury indictment 3. information filed by prosecution based on sworn complaint of vicitm
What does centi mean?
1/100
What does milli mean?
1/1000
What does deca mean?
10
w/in how many days of assignment to a position involving bloodborne pathogens must employers offer hep B vax?
10
How many STR loci can you get information from using Profiler and Cofiler kits?
10 loci
The National Research Council (NRC) recommends that the minimum number of people in the database should be at least how many?
100
What does hecta mean?
100
What does kilo mean?
1000
What OSHA standard is referred to as the "right to know" training?
1910.1200 Hazard Communication Standard (HCS); mandates employers must be informed of hazards of all chemicals used.
The Federal Bureau of Investigation (FBI) crime laboratory opened in what year?
1932
Single beam Instrument with Diode Array Detectors
Advantages: Data accumulation rapid, simple design, few moving parts, little calibration needed, minimal maintenance Disadvantage: instrument has limited resolution
Who was responsible for courts acceptance of documents as scientific evidence?
Albert Osborn
Most commonly used protein comes from a source other than human to avoid the antibodies recognizing the human albumin
Albumin
What is the most abundant protein in blood serum?
Albumin
_____ is credited with the first practical forensic application of human DNA analysis.
Alec Jeffreys
Saliva is alkaline or acidic?
Alkaline
Dragendorffs Reagent
Alkaloid base, Primary, secondary, tertiary amines
Dragendorff's reagent
Alkaloidal base, primary, secondary and tertiary amines Orange, Red-orange, brown-orange
During crime scene processing which type of evidence is collected first?
All fragile evidence
The active phase of hair growth is known as ____.
Anagen (active); Telogen is naturally shed hair and Catagen is attached but not actively growing
Which hair growth phase(s) can be used for DNA analysis?
Anagen and Catogen
What is the best stage for DNA comparison of hair?
Anagen stage
What are th 3 stages of growth?
Anagen, catagen, telogen
How long does each stage of growth last?
Anagen- up to 5 years Catagen- 2-3 weeks Telogen- 2-6 months
Criminalistics
Analyze and interpret physical evidence
Document Examiners
Analyze ink, paper, toner in questioned documents
Which presumptive test for blood reacts with blood proteins to give a light to medium blue color? A. Luminol B. Leucocrystal violet C. Amido Black D. Bluestar
C. Amido Black
The best screening test for saliva is ___ A. Acid phosphatase mapping B. P30 mapping C. Amylase mapping D. Glucose mapping
C. Amylase mapping
Which of the following tests has the potential to be the most harmful and therefore is not commonly used? A. Tetramethylbenzidine B. Phenolphthalein C. Benzidine D. Luminol
C. Benzidine
When using DNA analysis for species determination the DNA probe used is known as A. D40MER B. SWD17 C. D17Z1 D. SWMER17
C. D17Z1
The general principle of gel electrophoresis is to separate molecules based on A. Molecular weight B. Shape C. Electrical charge
C. Electrical charge
The ABO blood typing system was developed by whom? A. Ouchterlony B. Watson C. Landsteiner
C. Landsteiner
If genotype frequencies are one locus are independent of genotype frequencies at a second locus, the two locus are said to be in ____. A. Hardy-Weinberg equilibrium B. Likelihood equilibrium C. Linkage equilibrium D. Product equilibrium
C. Linkage equilibrium
What does the genescan software NOT do? A. Identify DNA fragments B. Quantitates DNA fragments C. Sizes DNA fragments D. Assigns allele calls to the peaks
C. Size fragments
mtDNA [can/cannot] be analyzed in cases of mass disaster using burned or highly degraded samples.
CAN
the complementation of two recombinant protein fragments that can spontaneously recombine to form a fully functional protein enzyme. The larger of the two fragments is referred to as the enzyme acceptor (EA) while the smaller fragment is termed the enzyme donor (ED). The activity of the fully functional enzyme is dependent on the ability of these two fragments to combine, and can be utilized in an immunoassay technique.
CEDIA cloned enzyme-donor immunoassay
Decide which statistical test is associated with the info for each: sample variance must be known, used in labs to deter critical parameters and their variability
CHi-Squared Distribution
The national DNA database is called
CODIS
A __________ is used to maintain a list of all personnel entering and existing a secured crime scene.
CS log security
These are 1 epithelial thick, fragile and blood can only flow single file.
Capillary veins
What is the difference between individual characteristics and class characteristics?
Exact identity compared to a common origin
• Dichroism and pleiochroism • Sign of absorption • Birefringence • Angle of extinction • Sign of elongation • Refractive index • Other microscopic parameters
Examination by polarized microscopy shows crystallographic features than may be used to identify specific compounds, such as:
List one of the several duties of a Forensic Pathologist.
Examine autopsy & the history of the person due to their death & witness testimony.
Process of removing DNA from dried stains includes: Chelex, organic and DNA IQ
Extraction
What part of the body is the fluid extracted from that is called vitreous humor?
Eye
T/F The Y-STR is found only on the X-chromosome.
False (Y on Y)
T/F Capillaries deliver oxygenated blood to the veins.
False (deliver O2 to tissues)
T/F Polymorphic enzymes have the same structural forms of an enzyme with the differing functional properties.
False (differing structure, similar function)
T/F ALS visualization of suspected semen stain with a crusty touch is considered positive
False - ALS is merely a screening tool
T/F, most forensic labs deal w/radioactive materiels
False - excepts are when radioimmunoassays are conducted
T/F regarding human subjects: there is no difference between fetuses, children, pregnant women and prisoners than men and women who are not potentially child-bearing
False - fetuses, children, pregnant women and prisoners are restricted more than others
T/F there several confirmatory tests for vaginal secretions
False - tests are presumptive in nature
T/F int he absence ofan appropriate SOP for a specific analyte in a spec. matrix, the lab is released from responsibility for preparing a SOP
False -- lab is responsible for the SOP
T/F Good quality management can result and be the cause of a nubmber of common laboratory errors
False --poor quality management
a less serious error failing to identify the presence of a drug when it is known to be present in the sample. associated with the failure of the analytical test and can result in critical data being lost.
False Negative
is identifying a sample as being positive for a particular drug when in fact none is present. associated with contamination of the case sample with the drug in question, or failure of the analytical test to produce an accurate result.
False Positive
T/F Degradation reduces peak height by causing an upward slope across the electropherogram?
False, degradation is seen as a downward slope
What type of scientific evidence is defined by "use of microscope comparisons of markings to associate an item of evidence with a particular source?"
Firearm Toolmark
Decide which statistical test is associated with the info for each: This is an exact test that is used to calculate the null hypothesis
Fisher's Exact probability test
Decide which statistical test is associated with the info for each: Post-hoc test. F-distribution is used to calculate the critical value
Fisher's Least Significant Difference test
Semen fluoresces due to the presence of _______.
Flavins
what are the two most commonly performed confirmatory tests in forensic science? they can be used to identify a compound to the exclusion of all others.
IR spectrophotometry and mass spectrometry
what's the diff b/t ITT protocol analysis and per protocol analysis?
ITT includes all test subjects regardless of randomized roup, even if they fail to follow protocol - it may mimic 'real life' to a better extent. per protocol includes data only of the test subjects who followed the protocol and thus the data may better indicate the true potential of a drug or treatment
When using either digital or film cameras at a crime scene the first image or frame should be of a __________.
Identifier
Components of Analytical Procedures
Includes: --extraction of the test compound from a matrix --construction of a standard curve --quantitation of the test article --quality control of the analytical process
List 4 routes of exposure to poisons at a crime scene.
Inhalation, Absorption into skin, Injection, Ingestion
What does observing a crime scene involve?
Initial walk through Plan of action
Fingerprints may be recorded using a live scan system or rolling the fingerprints in _______ onto a fingerprint card.
Ink
What type of board approves studies performed on human subjects?
Institutional review board (IRB)
How can the medulla be classified?
Interrupted, fragmented, continous, stacked, absent
Engineering
Investigate structural failure and investigate accidents
Weak acids and bases are incompletely or slightly ionized in solution. The extent of ionization is described by the equilibrium ionization constant, Ka.
Ionization Constants
The agglutination of RBC's with human sera was first observed by whom?
Landsteiner
What are the 5 requirements of Hardy-Weinburg Equilibrium?
Large population, No migration, No mutation, Random mating, Natural selection
This test must be done on fresh blood stains. It is used to test for presence of blood antibodies
Lattes test
A newborn baby of a mother Le(a-b+) and father Le(a+b-) will type as what?
Le(a-b-) [- is dominant]
If you are LEFT handed, the order of movement would be [Left to right/Right to left].
Left to right
RFLP is [equally/more/less] sensitive as PCR methods.
Less
What is a reason that urine is used most frequently in pre employment screening?
Less expensive, no puncture
Explain why the Locard Principle is important to crime scene work?
Links physical evidence from or to victim, subject and/or crime scene.
What is the SI unit for volume?
Liter
Duquenois-Levine Test
Marijuana, hashish - Violet Blue/Violet Brown Coffee - Violet Patchouli Oil - Green/Blue
what does duquenois-levine reagent test for and what is the final color?
Marijuana, hashish, thc, *violet-blue* and *violet brown* ; roasted coffee (violet); patchouli oil tea (green-blue)
Opium Alkaloids Final Color, Purple
Marquis reagent
What is the formula to calculate density?
Mass/Volume
Who should certify that the body is "dead"?
Medical examiner
Regarding MSDS: When are chemical manufacturers to provide privileged information as to the identity of proprietary compounds?
Medical or other emergencies, w/ understanding that the info will not be made available to anyone else.
This is blood spatter when the impact velocity is 5-15ft/sec, the spatters are 1-3mm and are typically a result of a beating, stabbing
Medium Velocity
If someone is beaten with a baseball bat, which type of SPATTER would you expect to occur? High velocity, Medium velocity, Low velocity
Medium velocity
What is the SI unit for length?
Meter
Mandelin Reagent
Methadone
Mandelin Reagent
Methadone Blue
What does mandelin reagent screen for and what color does it turn?
Methadone, *blue*
Is it possible for a document examiner to tell the personality of an individual from his or her handwriting?
No
Schedule I definition
No currently acceptable medical use; lack of accepted safety for use under medical supervision; high potential for abuse
Bodily Excretion and Polarity
Normal bodily excretion of drugs becomes polarized by the liver so that it can be eliminated in urine When analysts are trying to extract drugs they want to get the drug to be non-polar so they can extract it into the organic solvent phase
Describe the radial diffusion test for saliva
Not sensitive, false +s. uses agarose gel w/ starch, extract of unknown sample is added to hole in gel, diffuses - amylase in sample hydrolyzes starch produces a clear area after staining w/ iodine. Size of area proportional to amount of amylase in sample.
Pharmacokinetics of Opiates
Opiate receptors - analgesia, miosis, sedation/drowsiness, tolerance/dependence Mu and Delta receptors - Euphoria, nausea and vomiting Kappa receptors - dysphoria, diuresis
Marquis Reagent
Opium alkaloids
Which extraction method would be best suited for tissue samples?
Organic
DNA from gum and spongey bone are best extracted using which method?
Organic Extraction
Type of DNA extraction that uses a mixture of chemicals. Best for degraded or dirty samples
Organic Extraction
Which extraction methods is best for a buccal swab or reference standard?
Organic or Chelex
catalytic effects changing surface pH direct reaction with the drug.
Other compounds present in the drug sample can affect the stability of the drug as a result of
The most common species determination test was developed by whom?
Ouchterlony
Photochemical reactions occur when a molecule absorbs a photon of light energy to produce an unstable excited state molecule. This extra energy can be lost by several different mechanisms: • Fluorescence • Physical decay (loss of energy in the form of heat) • Chemical decay (breakdown of the drug) Ultraviolet light is the most energetic light in the spectrum and can induce chemical decay. Many steroid drugs are susceptible to photolysis.
Photolysis
• Changes in the crystal form of a drug • Aggregation of drug particles in a suspension • Loss of volatile chemical compounds to the atmosphere and container (alcohol)
Physical Decomposition
- Traces of impurities in street drugs may cause misleading results or interfere with crystal formation making identification difficult - Impurities present may provide an unsuccessful test - Considerable training and experience in chemical microscopy is needed to identify drugs based on the crystal form of the products of specific test reagent reactions - The technique possesses some degree of subjectivity - Crystals of one type may show a wide variation in form - One substance with one reagent may give more than one type of crystal product
Pitfalls of microcrystal tests
CEDIA pros & cons
Pros - less steps, easy to automate, long shelf life, reduced interference, direct linear response over large range of concentrations Cons - potential matrix interferences, potential false negatives
KIMS pros & cons
Pros - less steps, relatively inexpensive, microparticle conjugates more stable than enzyme drug conjugates, long shelf life, interferences tend to cause false positives, reduced interference due to measurement over time Cons - extensive instrument maintenance due to the microparticle conjugates sticking to tubing, a smaller linear range than CEDIA or EMIT
ELISA pros & cons
Pros - maximize sensitivity, low sample vol., long shelf life, oriented antibody, optimal enzyme conjugates Cons - requires many wash steps, sodium azide inhibits horseradish peroxidase, supplies are not reusable
RIA pros & cons
Pros - maximizes sensitivity, minimal matrix effects, rare false positives Cons - requires many wash steps, difficult to dispose radioactive waste, short shelf life
What type of sampling is required for scientific evidence?
Random, non-biased
Erythrocytes are also known as ____.
Red Blood Cells (RBC's)
What is the main difference involving the cartridge cases between a revolver and a semiautomatic firearm.
Revolver pellets stay in the gun, but the semiautomatic case eject from the gun.
List ONE of the three changes of decomposition.
Rigor, Liver, Algor
Endogenous compounds present in the sample matrix, as well as analytes in which we have no interest can then be removed by washing the extraction cartridge with a suitable solvent. Any combination of aqueous and/or organic solvents can be used for this purpose, the goal being to remove as much of the interference as possible without eluting the analyte of interest.
Rinse (Wash) (TLC)
What is the difference between routine and preventative maintenance?
Routine- includes cleaning, clearing hoses Preventative-changing out old parts and such
Difference b/t routine and prev.maintenance?
Routine: procedures to protect equipment and/or personnel- rinsing HPLC, replacing GC Septa. Preventive: anticipates probs cannot be handled using routine maintenance - oil replaced, internal parts cleaned, worn parts replaced before they break/damage, etc.
Properties of Common Stationary Phases
Silica gel--weakly acidic, very popular due to hardness and resilience Cellulose--uses partition process for separation, 50um spherical particles optimal, run slower than silica plates of same thickness Ion Exchange Resins--ctg quaternary ammonium or sulfonic acid groups for ion exchange bonded to surface of the plate, good for high MW substances and amphoteric compounds, mobile phase is usually a strong acid or base
What type of medullas do animals usually have?
Stacked, continuous or interrupted
Factors that affect reproducibility for TLC
Stationary phase --quality of the binder or absorbent, impurities or diff in thickness of the phase on plate Mobile Phase--impurities in the solvent--prepare daily Develop Tank--ill fitting lid, unequilibrated solvent vapors Temp--solvent runs faster when warm, volatile substances evap faster and Rf values tend to decrease Develop Distance--increased Rf with increase in running distance due to evap of solvent Mass of sample--increased mass will increase Rf
Radio-immunoassay (RIA)
Strengths: sensitive, strong assay response, resistant to matrix effects, bile and blood, amphetamines, barbiturates, opiates, PCP, cannabinoids, benzos, LSD, for ricin in urine and THC in hair Weakness: radioactive waste, difficult to automate
Describe one advantage and disadvantage to using hemastix
Strips are a one step process and can be done quickly without reagents. Specificity is lower than other test and can be difficult to get stain to transfer.
the solvent, and any impurities present in the test sample will also be concentrated and may hinder or distort crystal development.
Supersaturation may occur as a result of evaporation
List an examination that is conducted during an autopsy.
T-shape when they go in the stomach & chest.
what are the 2 microchemical confirmatory test for blood?
Takayama (hemochromogen) and Teichman reaction
One of the most common confirmatory tests for blood is called ____.
Takayama Test
Name 2 confirmatory tests for blood.
Takayama and Teichman
Describe anthropometry.
Taking body measurements as a way to distinguish one individual from another
T/F Blood groups agglutinate with their same antisera. Ex. A blood agglutinates with antisera A. All except O do this.
True
If the refractive index of glass matches that of a liquid, what will you see when ou put the glass sample inside the liquid?
The glass will disappear
What is the Daubert standard?
The stringent review by the court of practices applied to evidence examination to determine admissibility. Judges function as the gatekeepers.
Securing the crime scene is the responsibility of ____.
The initial responding officer
The range of values for [H+] in aqueous solutions at room temperature (20-30°C) is from 100 - 10-14. Therefore the pH scale is from 0-14. Note that if an acid is added to water pH < 7 if a base is added to water pH > 7 The pH scale depends on the temperature of the solution for which the pH is being measured. pH + pOH = 14
The pH Scale
What is the Frye hearing?
The presentation of generally accepted [scientific] practices to the court for admissibility
What is a "Likelihood Ratio"?
The probability representing DNA from the evidence sample and that from the suspect are the same to the probability of a match if they came from different people.
T/F Blue eyecolor written as Bb is a genotype
True
extremely variable accelerated by the presence of water and/or heat (known to occur with some steroids and antibiotics).
The rate of transformation of one form to another
why might Leucomalachite Green be preferred in a presumptive test for blood over the Kastle-Meyer test?
The reagent phenolphthalein is time consuming to make
What specific piece of equipment is needed to process PCR?
Thermal Cycler
What organism is Taq Polymerase found?
Thermus aquaticus
Elements of an SOP "in house"
Title of SOP, author, issue date, ID #, Page x of y, approval signature (senior lab official), some subsections include: Scope--elaborates on title Referenced Docs--source providing info for the SOP (federal standards, journal articles, SOP's of other labs) Defiinitions/Terminology--unfamiliar terms, common abbreviations Summary of Methods--briefly describes equip and general procedures Inferences--compounds that will interfere with results are noted Apparatus and Materials--lists devices, columns, detectors used in analysis, also should include syringes, bottles, vials Reagents--solvents, stock standards, reference standards, calibration standards, internal standards, surrogates, include conc. hazards, chemical grade, storage conditions Sample Collection, Preservation, and Handling--specific methods for handling a sample Procedures--Instrument or method calibration, sample prep and analysis, troubleshooting, calculations Quality Control--Procedures est. to demonstrate that instrument calibration, sample prep and analysis are preformed correctly. Method Performance--The min detectable level IMDL) of a substance in diff. matrices, % of recovered chemical Miscellaneous--flow charts, diagrams
What is identification used for?
To determine physical or chemical identity
What is comparison analysis used for?
To determine whether a common origin exists
DNA fragments will travel [toward/away] from the anode.
Toward the anode; away from the cathode
The study of poisons refers to what?
Toxicology
What category of stain would a bloody shoeprint classify as? Expirated stain, cast-off stain, transfer stain projected stain
Transfer Stain
List a crime scene measurement method.
Triangular search and Baseline
T/F A questioned or crime scene sample that is identical to a known standard at any given number of loci is referred to as a match.
True
T/F ABO antigens are found in plasma, semen and saliva
True
T/F According to the Lewis Grouping System, newborns and 20-25% of blacks are (Le a-b-)
True
T/F Blue eyes are a phenotype
True
Inferential Statistics (inductive statistics)
Type of statistics where probability theory is utilized to determine if results are significant, clinical studies are prone to this bc there are so many variables associated with humans (such as--genetics, life style and exercise, environment, medications, indiv. not following protocol), control groups are utilized to make comparisons when there are less than 30 samples (n< 30) the distribution is less likely to be normal Student's t-distribution can be used to deter the confidence interval for both small and large populations, best when examining paired averages--where one wants to know whether the avg values are diff for a specific parameter obtained by two diff methods Duncan's Separation of Means Procedure--way to statistically deter whether more than two averages are statistically different CHi-square test--used to deter whether two groups of data are different, measures diff bt observed and theoretical frequencies of an event occurring
Describe the proper way to collect and store the evidence below: Fibers
Use tweezers to pick up fibers after photographing. Put into small envelope, seal, then place in a larger envelope and label.
Describe the proper way to collect and store the evidence below: Hair samples
Use tweezers to pick up hairs after photographing. Put into paper bindles, then place in a larger envelope and label.
Which one of the following methods is not a DNA quantitation technique? Slot blot, Western blot, Real-time PCR, Picogreen Assay
Western Blot
What evidence cannot be packaged immediately? Why?
Wet evidence because it could be destroyed by mold or mildew
• Weight if in powder form (note if dry weight or wet weight) • Amount if in tablet form • Number of packages • Nature of the substance • Color • Odor (if obvious) • Condition- wet or dry
When samples are acquired for testing the appearance and features of the substance must be recorded. These features include:
A Forensic Scientist working with Controlled Substances has many duties provide 2 duties that were discussed in class.
You must photograph and document. Weigh the package then package the substance. Send to the lab to test the controlled substance.
Complete satisfaction of Hardy-Weinburg Equilibrium would suggest that THETA = ___?
Zero.
Forensic Court reports covered by FBI Quality assurance standards for Forensic DNA Testing Labs should include
a case identifier, description of the evidence examined, a description of methodology, the locus used, results/conclusions, interpretive statement, date issued, disposition of evidence, a signature and title in addition, ISO17025 requires that: total # of pages on any hard copies of the report, the name and address of the client, unique ID # on report, descrip of the sampling plan, date of receipt of test item
Calibration
a comparison of a measurement, standard, instrument or item with a standard instrument of higher accuracy to detect and quantify inaccuracies and to report or eliminate those in inaccuracies by making adjustments. (physical measurements vs. acceptable standards) Should cover the entire operating range of the instrument, 3 or more points used, numerical data in SI units.
base
a compound that accepts hydrogen ions
acid
a compound that donates hydrogen ions
in QA/WC work, what is a study protocol?
a document that lays out the objectives and a detailed description of the total experiment or research activity
First Pass Metabolism
a drug is absorbed across the gut wall, taken to the liver, some metabolism may occur before the drug enters the systemic circulation
drug
a natural or synthetic substance that is used to produce physiological or psychological effects
regarding developmental validation, what is needed in order for independence studies and allele frequency determinations to be made?
a number of samples from a population
field sobriety
a preliminary test performed to ascertain the degree of a suspect's physical impairment and whether further tests are justified
field sobriety test
a preliminary test, usually before breathalyzer test, to assess balance, physical impairment and attention level.
High Performance Liquid Chromatography (HPLC)
a sample is injected onto the column through an injection port using a syringe. Ind. components are separated on the column are detected as they emerge by a detector. Components---- Eluent--this can be water, buffer soltn, organic mixtures. HPLC grade liquids should be used to minimize the presence of impurities. They should be filtered and degassed. Effect of pH--you want to avoid ionization of the compound. Adding a buffer at the required pH usually prevents ionization. For acids, buffer pH=3 added to mobile phase, for bases, buffer of greater than pH 8 should not be used bc this will deteriorate the column. Therefore an ion pair agent is employed and the pH of the mobile phase is kept acidic. ex. sulfonic acid and N+ Pump--this delivers the eluent to the column at a controlled rate and higher pressure. Flow rates are typically .5-2 ml/min. The flow rate must be continuous. Injection system-uses a fixed loop that is filled with excess sample and holds a precise amount of soltn int the injection column. Columns--There are two types of columns 1)absorption or normal phase- adsorbent material is used, usually silica, solutes bind to the column based on solubilities-polar compounds have long retention times, works best for non-polar substances 2) partition or reverse phase-in these polar compounds elute first, and non-polar compounds are strongly bound Detectors-- UV-Vis, Dioade array, Fluorescence, Electrochemical, refractive index, mass spectrometer
immunogen
a substance injected into an animal, causing production of antibodies to the injected substance
poison
a substance that through its chemical action usually kills, injures, or impairs an organism
horizontal-gaze nystagmus
a type of field sobriety test to assess vision capability
the point of origin of a fire can often be determined by examination of burn patters in what pattern?
a vee shape
If electricity and ventilation fails, what must sealed systems such as cold or warm rooms have?
a way for employees to rapidly exit
Standard Operating Procedure (SOP) defined
a written document that details the method for an operation, analysis, or action with thoroughly prescribed techniques and steps that is officially approved as the method for performing certain routine or repetitive tasks
metabolite
altered form of the compound for excretion purposes
natural variation in a person's handwriting is ___ when compared to the variation between individuals
always less
Examples of Drugs that are acids
amobarbital, aspirin, LSD, IBU, penicillins
Process that produces millions of copies of DNA using PCR
amplification
indented writings on evidence can be visualized with an instrument called
an Electrostatic Detection Apparatus (ESDA)
a flammable liquid that was used to state a fire is called
an accelerant
Liquid-Liquid Extraction
an immiscible organic solvent is added to an aq soltn of a drug, this will cause a partition bt the two phases. A partition coefficient can be calculated from the conc of the drug in the organic phase divided by the conc in the aq phase.
which is not a growth phase of hair
anagen
narcotic drugs are
analgesics
Screen
analysis that is employed to ID/separate various classes of analytes within a sample ex. drinking water using GC/MS
the ACE-V method of print identification involves
analysis, comparison, evaluation, and verification
What are the 3 differences bt human and animal hair?
animals have a patterned medulla, wider medulla and inconsistent color
Preventive Maintenance
anticipates problems that cannot be prevented with routine maintenance ex. replacing oil, cleaning internal parts, replacing worn parts
antigen
any substance that stimulates an animal lymphocyte to produce an antibody that specifically binds it
forensic toxicology
application of toxicology in the law
40
approximately ___% of traffic deaths in the US are alcohol related
what is the exception to the "1 species per room" housing rule for animals?
aquatic animals - each species maintained in separate tanks,same room
the basic fingerprint class patterns are:
arches, loops, and whorls
Mechanisms of Drug Degradation
chemical instability of drugs can be attributed to hydrolysis, oxidation or photochemical changes Hydrolysis--this is the cleavage of chemical bonds by the addition of water, esters and amides do this, ex. cocaine, procaine Photolysis--rxns that occur when a molecule absorbs a photon of light energy to produce an unstable excited state molecule--extra energy can be lost via fluorescence, physical decay, chemical decay, UV light is the most energetic light in the spectrum that can cause chemical decay, many steroid drugs are susceptible to photolysis
name 3 types of specialized waste laboratories must dispose of
chemical, biological, radioactive
Chemical Exposure (storage)
chemicals should not be exposed to direct sunlight, heat source, or stored under sink, bottles should be placed below eye-level, and in cabinets with see-through doors. Flammable and hazardous chemicals should be stored according to their labels. Separate chemicals that could react together. Gas cylinders should be stored outside the laboratory when not in use. Cylinders must be stored upright with their valve protection caps on. Chemicals should be stored separate from chemical waste.
Rf values
chromatographic measurement that describes the distance the substance travels from the origin divided by the distance the solvent front travels. If the substance is a spot you measure from the middle of the spot, but if the spot has a tail then you measure from the densest area of the tail. Rf = distance the substance travels from the origin/ distance the solvent travels from origin
Assay Techniques: CEDIA
cloned enzyme-donor immunoassay is based on the complementation of two recombinant protein fragments that can recombine to form a fully functional protein enzyme. There is an enzyme acceptor (EA) and an enzyme donor (ED). The assay components have a free floating antibody that recognizes the drug of interest.
The smaller the diameter and denser the pattern means that gun was ______ to the victim
closer
Additive interactions
combined effect equals sum of each effect (e.g. 1+1= 2)
Synergistic interaction
combined effect greater than sum of each other (e.g. 1+1= 3)
Bilirubin
comes from break down of red blood cells, excreted into bile (used to excrete lipids and lipid soluble drugs), impaired bilirubin leads to jaundice
placing a fixed quantity of the antibody in a tube together with a fixed quantity of labeled drug and the drug sample to be assayed. The specific binding sites on the antibody bind both the labeled drug molecules (*Drug), and the unlabeled drug molecules present in the test sample.
competitive binding assay
base
compound that accepts hydrogen ions
acid
compound that donates hydrogen ions
Specific Absorbance (A)
conc expressed as g/100ml
Phase 2 metabolism
conjugation reactions, usually with glucuronic acid (takes a drug and makes it bigger by and more polar) some drugs may enter directly into phase 2 if already polar enough
Laws of Absorption: Lamberts Law
considers instrument factors, states that for a given conc (C) the radiation transmitted through a homogeneously absorbing system, transmitted intensity (I) decreases exponentionally with increasing path length (width of the sample cell)
Laws of Absorption: Beers Law
considers sample conc, for a given path length the transmitted intensity decreases exponentially with increasing sample concentration of a homogeneously absorbing system.
Chain of Custody
defined as an unbroken trail of accountability that ensures the physical security of samples, data and records, signature or initials of each individual receiving or transferring the evidence should be recorded with an identifier #, should include the date and qty of the batch Prevents 2 common methods by which analytical results can be challenged: 1. showing that the analytical procedure was not accurate 2. raising doubt about whether or not this was the correct sample Legally defendable when it can be shown that the original sample could not have been exchanged for other sample
Ionizing Radiation
defined as radiation from alpha, beta, gamma and x-rays, high speed electrons or protons, neutrons and atomic particles. UV, IR , Vis, sound or radio sources are NOT considered ionizing radiation. Radiation is strictly regulated by federal and state nuclear regulatory commissions. Electron capture gas chromatography detector, a stainless steel cylinder ctg the beta-emitting radionuclide nickel-63, are exempt from regulation.
what type of water is required for post-amplified side of the DNA lab?
deionized
Partition Coefficient (K)
dependent of the nature of the drug and the organic solvent, temperature, and degree of ionization the partition bt the organic and aq phases depends of hydrophilic (water liking) or lipophilic (fat liking) nature of the compound. Polar compounds partition into the aq phase Non-polar compounds partition into the organic solvent phase
Stability
dependent on the type of procedure under study, parameters that deter the stability of the analyte include: pH, tem, humidity. Stability of extracted biological samples is deter during assay develop/validation by replicate injections of the sample over 24 hr period or longer
labs performing routine analyses such as pesticide lvls in soil, creatinine lvls inblood, presence of controlled substances in saple, etc, usually use which type of statistics? (descriptive or inferential)
descriptive
name the components of a study protocol
descriptive title, study objective, ID of test and control articles, sponsor,testing facilities, other participants and justification of the study, detailed description of experiment, experimental design, start/finishdates, approval, statistical methods used, appendicies, amendments, deviations
Describe the method used in the Jaffe test
detects creatinine. Creatinine forms red compound with addition of piric acid
Standard Curve
deter by calibrators by a linear least squares fit to the equation y = mx +b x = conc and y = ratio of drug peak area
Quantitation Limit (LOQ)
deter by reference to signal to noise ratio and establishing a min conc at which the analyte can be reliably quantified. A typical signal to noise ratio is 10:1. The quantitation limit and method used to deter LOQ should be included in the validation report
precision assessment
deter from the slope of the replicate std curves on the same day and over a 2 week period, y-int < 25% of the value of the standard curve calibrator
post-mortem forensic toxicology
determines the absence of presence of drugs and their metabolites, chemicals such as ethanol and other volatile substances, carbon monoxide and other gases, metals, and other toxic chemicals in human fluids and tissues, and evaluates their role as a determinant or contributory factor in the cause and manner of death
human-performance forensic toxicology
determines the absence or presence of ethanol and other drugs and chemicals in blood, breath or other appropriate specimen(s), and evaluates their role in modifying human performance or behavior
Assay Techniques: FPIA
fluorescent polarization immunoassay uses the diff in rotation of bound and free fluorophore-labeled drug. Abs light at 485nm and emits light at 525nm. Polarized light that impinges on a flurosin-labeled molecule produces emission of plane-polarized light. Binding of the molecule to the antibody reduces the amount of rotation which increases the intensity of the polarized light Disadvantages: possible background interference for other drugs, enzyme labels could be inhibited by endogeneous molecules, fluorescent enhancement
two separate storage areas for animals are needed for ____ and ____. they cannot be the same areas used to store or analyze test materials
food and bedding
Antigens
foreign body, molecules that elicit an immune response and bind with antibodies
unknown fingerprints are entered into the data base as:
forensic files
number of calibrators to construct a standard curve? depending on what?
four to seven, range of concentrations to be quantified
What part does a microscopy analyst look for to ID a marijuana plant?
glandular trichomes
Internal Validation
has to be conducted by a laboratory when develop. validation has been performed by another laboratory or at another site of a multi-site laboratory Must be sufficient to show the reliability of the technique as practiced by that laboratory FBI Quality assurance standards for forensic DNA 1. accuracy, precision, reproducibility 2. locus must be defined and characterized 3. species specificity 4. sensitivity, stability and mixture studies 5. database and independence testing 6. allele and frequency distributions determined 7. non-probative sample study 8. match criteria established
what happened in the steven lawrence murder that generally cannot occur in the United States
he was tried again for the same crime
The Takayama confirmatory blood test works by ______ reacting with _______ to produce red feathery crystals that can be examined under a microscope.
hemaglobin, pyridine
Cloned Enzyme Donor Immunoassay (CEDIA)
homogeneous (directly proportional) - genetically engineered enzyme, 2 enzyme fragments EA + ED (drug labeled), CPR is produced when fragments bind, with more drus in the sample, the free drug binds to the antibody, so the fragments bind together producing CPR and galactose, MORE drug = MORE CPR produced
Enzyme Multiplied Immunoassay Technique (EMIT)
homogeneous (directly proportional) - opposite of ELISA - enzyme labeled drug when bound to antibody, activity is reduced, substrate is oxidized, enzyme activity measured by absorbance. MORE drug = MORE enzymes NOT bound to antibody, substrate can bind to enzyme
quantitation
interpretation purposes, level of drug within the body (therapeutic v. lethal drug), use blood, internal standards are used
Phase 1 metabolism
introduce/unmask functional groups by way of alkylations, hydrolysis, acetylation, and reduction (e.g. adding acetyl group, or hydroxyl group) enzymatic transformation of functional groups purpose - to convert lipophilic molecules into more polar molecules
Routine Maintenance
involves simple procedures with the purpose of protecting the equipment and/or personnel ex. replacing GC sept, rinsing an HPLC system prior to shutdown to prevent build up, disinfecting biological safety cabinet after each use, cleaning pump block on the genetic analzyer
Auxochrome
is a group of atoms attached to a chromophore which modifies the ability of that chromophore to absorb light.
Validation of Analytical Procedures
key parameters in the validation process are: 1. purity and identity of the neat test sample 2. homogeneity of soltns 3. extraction recovery 4. range of standard curve and control samples 5. linearity and precision of standard curves 6. sensitivity 7. specificity 8. accuracy and precision 9. stability 10. quality control
MSDS (Material Safety Data Sheets)
labeling on the ctgr of a chemical that includes hazard information about the product, should provide the following info on the label: --Product ID-name, synonyms, formula, MW and CAS # --Composition Info on ingredients --Health hazards/first aid/personal protection --Handling and Storage --Flammability --Physical and chemical properties --Toxicology --Disposal Methods --Transportation --Regulatory --Other Limitations to MSDS-- 1. Manufacturer is not required to perform exhaustive studies of health hazards and toxicology, etc of every chemical sold but must report what is known about a compound 2. The composition of some chemicals is sometimes considered proprietary knowledge. In the case of a medical or other emergency, the manufacturer should provide privileged info as to the ID of the compound to the appropriate medical personnel
what does LIMS stand for and what is it used for?
laboratory information management system - track samples, test results, scheduling, etc.
the majority of DNA is
non-coding
Fluorescence Spectrophotometry
measures light that emitted from molecules that are excited to higher energy levels after the abs of light, molecular structure and chemical enviro will deter whether a substance will fluoresce. Compounds should have flat rigid structures with high concentrations of delocalized e- Advantages--more sensitive and more specific than UV-Vis, but not all drugs will fluoresce. Disadvantages--Limited # of drugs will fluoresce, tough to distinguish bt parent drug and metabolite Examples of drugs that naturally fluoresce: --Chlorquine, Dantroline, LSD, Propanolol, Quindine
pH scale
measures the strength of acids and bases
excretion
mechanism of elimination that includes breathing and urine formation
oxidation
mechanism of elimination that occurs in the liver
role of forensic pathologist
medical examiner, review case history and information from scene investigator, perform postmortem examination, collect appropriate specimens, gather data including toxicology, determine cause and manner of death, give family answers
A baseball bat beating involves _______velocity blood spatter.
medium
Official SOP's
methods outlined for many routinely analyzed compounds that can be Environmental, Clinical and Pharmaceutical, Manufacturing Prior to SOP use, the accuracy, precision, selectivity, reproducibility, and repeatbility of a method must be demonstrated before it is actually used
what is the most reliable technique for confirming the presence of sperm?
microscopic analysis
2 out of 3 of the following tests can identify marijuana
microscopy, color test or TLC
photographs of crime secenes must include overall views, close ups, and _______?
midrange
molecular label
might be a totally separate molecule from the antigen, or a modification of it
how many concentrations is recommended to establish linearity between concentration and response?
minimum of five
protein precipitation
mix matrix with internal standard, then add organic solvent, proteins fall out of solution, may be centrifuged
Chromophore
molecular group that is responsible for energy absorption, usually conjugated systems with delocalized e-
blood
most important specimen for collection, collected in all postmortem cases, stay away from CENTRAL blood (stomach, liver, kidneys - high concentrations)
Thin Layer Chromatography (TLC)
most widely used technique for separation and ID of drugs, popular bc simple, cheap, reliable and selective, run several samples at a time, any sample dried on the plate will remain there uses mobile phase moving across a thin, uniform layer of finely divided stationary phase by capillary action, drugs migrate up the plate at diff rates due to: drug solubility in the mobile phase, drugs H bonding capabilities, degree of drug ionization (pKa) disadvantages---lower sensitivity and resolution than GC and HPLC
Interference
natural constituents of samples that can be co-extracted during residue analysis and need to be recognized, ex. Br in veggies, S is soil, CS2 in crops
used to rule out a drug or drug intoxication. the color produced by reaction with a reagent may be specific for a particular drug, but usually the color change is indicative of a class or classes of drugs (hence they are used solely as presumptive tests).
negative result for a color test
Physical Decomposition
no change in covalent bonds of drug, but involves changes to the crystal lattice structure. Chemicals with crystal lattice structures have narrow melting points. Amorphous solids when there is no such arrangement or structure have a melting point range of 10C. They also melt at lower temps that ones with a crystal lattice. Polymorphic Drugs--have more than one form of crystalline structure, different solubilities
antagonists
no pharmacological activity, blocks receptor sites, causes inhibitory effects
the fire triangle includes heat, fuel, and _____?
oxygen
What types of parameters affect stability?
pH, temperature, humidity
Validation of Analytical Procedures: Stability
parameters that should be heeded should be included in the report such as pH, temp, humidity. Stability of a standard curve, control and subject samples is prospectively demonstrated for the sample collection and storage conditions used for the study biological samples need to be validated with replicate injections of the sample over 24 hours
Pain management
patient's compliance with medication, polysubstance abuse, chronic pain (therapy, exercise, massage), testing frequency, specimen selection
analysis of samples should show a single major (peak, trough) in the chromatogram with retention time corresponding to that of the previously injected (standard, sample)
peak, standard
what do most presumptive tests for blood rely on?
peroxidase activity of hemoglobin and oxidation of chemical indicators
A circular pattern of GSR around the bullet hole means that the weapon was _______ the target.
perpendicular
this type of fingerprint is a three dimensional impression
plastic
What things should you note about the human remains at a homicide scene? (List at least 5)
position of body, any possible wounds, signs of lividity or rigor mortis, insects present?, condition of clothing
Applications of Immunoassays
postmortem investigations, workplace drug testing, human performance investigation (steroids) Typically an analysis is done on urine bc it has fewer interfering compounds and decomposition products. Blood assays are more complicated due to interfering compounds. Proteins are usually ppt before analysis using acetonitrile to eliminate interfering substances.
the rough sketch that is usually prepared at a crime scene is used to
precisely locate evidence
(accuracy,precision) is expressed as relative standard deviation of the intraday and interday replicate analysis for each control sample
precision
types of crime sketches include
preliminary, cross projection, smooth
Single Beam Instrument with a diode array detector
radiation is focused on the sample then passes through a monochromator with a fixed diffraction grating. The dispersed radiation is reflected onto a photodiode transducer that is composed of hundreds of photodiode array transducers mounted on a silicon chip. The slit width of the monochromator is adjusted to the width of one of the diodes so the output of each diode corresponds to radiation of a diff wavelength, by scanning these outputs sequentially a spectrum can be obtained. Advantages: quick data accumulation, simple design, few moving parts, little calibration needed, minimal maitenance required Disadvantages: limited resolution 1-2 nm, low specificity, therapeutic drugs are often too low to detect and metabolites can retain the chromophore which can contribute to the absorbance spectrum. This is why this test is only a screening test NOT a confirmatory test. Forensic Science applications: several drug classes have good UV abs, including: barbituates (phenobarbital), benzodiazepines (diazepam), phenothiazines, tricyclic antidepressants
what are the measures of precision?
range, standarddeviation, variance,relativestandard deviation
inhalation
rapid delivery to other target tissues, quick onset of effects, abuse, avoids first pass
intravenous
rapid onset, avoid solubility issues, avoid first pass metabolism, difficult to administer, painful needles, immediate effects
sublingual
rapid onset, convenient, poor bioavailability, primarily for low dose/ high potency drugs, needs to be lipophilic to cross membranes
What are some drawbacks to using Luminol as a presumptive test for blood?
reacts with Fe, Cu, Co, K, bleach, and plant peroxidase, causes loss of genetic markers, can dilute stains, wash away impressions
In a Christmas tree stain, what color are the heads of the sperm that contain the DNA?
red
Equipment
refers to freezers that store samples, thermometers, thermal cyclers, automatic pipettes, hood ventilation system, also items that measure some parameters are called more specifically instruments
postmortem redistribution
refers to the changes that occur in drug concentrations after death. It involves the redistribution of drugs into blood from solid organs
elimination
removal of a drug from the body in either unchanged form or as metabolites
Specificity for Chromatographic procedures
representative chromatographs should be used to demonstrate specificity. Resolution of the analyte peak from any other compound verifies specificity.
wet chemistry laboratory
requires sinks, bench tops, electrical outlets, and hoods. Some chemicals can be stored within the lab in a special cabinet. Separate cabinets are needed for acids +bases, volatile liquids, etc. Be sure not to store incompatible chemicals side by side.
Study Protocol: Start and Finish Dates for Study
retention of all experimental records is necessary, this includes hand-written observation, raw data obtained, protocols, data, documentation, reports an even specimens must be saved for some time (non-clinical study w/ application= 5, no application = 2 yrs) -this is mandated by the FDA EPA requires 2-5 yrs retention of record based on whether or not the results of the study are submitted to the EPA, clinical drug regulations state that records must be retained for two years after marketing approval or notification that the study was discontinued Protocol Approval - signed and dated by the sponsor and study director. Also signatures by quality assurance officers, consultants and others Statistical Methods-statistical methods should be summarized and rationale for their use included. Appendices--Ex. of the Informed Consent Form, SOP's, Ex of Owner consent form (animal), descript of collection sites, descript of special equip, validation studies, investigator qualifications
What substances gives false positives for presumptive blood tests?
rust, horseradish, vegatables
Transport of Test article tracking
sample must be maintained under appropriate conditions during transport from the field to laboratory, when biological liquid samples are involved they should be placed in a water proof closure inside the sealed shipping container, chain of custody (COC) should also be included during transport that states: name and address of the requesting company, list of samples with ID #, type of sample, manner of collection, test (s) requested, name and address of testing laboratory, signature of person delivering sample, signature of person receiving the sample. Shipping ctgr should be sealed or locked. Save all packaging and inspect for evidence of tampering.
Sample Contamination
samples must be separated from each other, and other sources of contamination, during transit, storage and analysis. Of utmost importance when dealing with volatile compounds and concentrates. examples of samples commonly contaminated during analysis--- rubber and plastic items--lubricating oils in gloves, CS2 from handling veggies/fruit drinking water--disinfection by-products
These are small drops that surround a parent stain
satellite or secondary spatter
what compound gives feces its characteristic odor?
skatole
the ______ the droplet, the ______the force that was used to create the spatter
smaller, the greater
Standards Solutions
soltns of drugs or chemicals of known concentration
Infrared (IR) Spectrophotometry
study of reflected, absorbed, or transmitted ER of wavelength 0.8um to 500 um. The IR spectrum is divided into three distinct regions--near IR = 12,500cm to 4000com, mid IR = 4000 to 400cm (the mid region provides useful info on pesticides and drugs), far IR 400-20cm based on the principle that molecules have specific internal vibration frequencies, the total energy of a molecule = the sum of it's vibrations When IR energy hits a molecule it causes energy transitions which gives rise to the abs characteristic of the compound. This only takes place when there is a change in the dipole moment of a molecule so most compounds with covalent bonds can abs. IR, but diatomic molecules do not. A sample inserted in an IR beam will absorb radiation at frequencies corresponding to the molecular vibration frequencies of the molecule and transmit all others. Can be used qualitatively and quantitatively-magnitude of abs of a chemical species is proportional to the amount of the chemical species present
Validation of Analytical Procedures: Range of Std. Curve and Control Samples
sufficient vol of the std curve and control samples should be prepared in a drug free pooled serum (or other bio matrix) for use of the remainder of the study. Std curve calibrators are prepared for each compound of interest in each biological matrix to be analzyed during the study.
Purpose of a final report
summarizes the work, provides interested parties information, documents that the laboratory has accomplished the task that it was funded, provide a record to the court of the results of forensic examinations
Wet blood stains should be collected by
swabbing
Describe how the BCIP test works
swabbing of stain and adding to tube w/ BCIP. If AP is present it will hydrolyze the BCIP creating blue color changes w/in 15 mins.
Validation of Analytical Procedures: homogeneity of soltns
taking aliquots from the top, middle and bottom of a sample ctgr. Concentrations should fall in an acceptable range.
who has overall responsibility for technical operations of the lab?
technical manager
mass spectrometry
technique based of the detection of vaporized molecules and their ionized (charged) fragments; detection and display of the spectra are based on the mass-to-mass charge ratios of the ions; method is specific for qualitative analysis and useful for quantitative analysis Advantages: specific, sensitive, qualitative and quantitative, compatible with separation Disadvantages: expensive, high maintenance, skilled analysts, not as sensitive as fluorescence, IF, more suited to low MW compounds, interfaces with liquid is hard
Immunoassays
techniques used to measure the presence and amounts of chemical substance in bodily fluids such as blood, plasma, serum, virteous humor and urine, work on the principle of competition bt a drug in the sample and a labeled drug of antibody specific for the drug of interest. The higher the conc. of the drug in the sample, the lower the conc. of labeled drug that will bind to the antibody. These tests need to be specific and sensitive.
competitive binding
the analyte and the labeled analyte competing for the binding sites on the antibody
who is responsible for the ID, maintenance and use of all records?
the archivist
Electronic Records
these records can be stored on a variety of media from 3" disks to CDs to computers. Limited access must be given for info stored on computers to prevent alterations and deleting info., 2 copies should be kept and stored at different locations STORAGE requirements all storage media have finite lifetimes, choice of storage media depends on--amount of data the medium can hold, ease of use, reliability, depends a specific operating systems, period of time the records need to be saved
Physiochemical factors
things that affect drug decomp rates include temp, catalysts, pH, nature of solvents extra drug adulterants can also affect the stability of the drug
Demonstrating specificity of assay when impurity or degredation product stds are unavailable
this is achieved by comparing the test results of samples ctg impurities or degredation products to a second validated method. This should include samples stored under relevant conditions of heat, light, humidity, acid/base hydrolysis, oxidation
Validation of Analytical Procedures: Extraction Recovery
this is deter by the comparison of the peak area observed with a soltn prepared at the same conc in the appropriate matrix, it is then extracted and injected into the chromatographic system. Three concentrations should be used.
Validation of Analytical Procedures: Quantitation Limit
this is done by the analysis of samples with known concentrations of analyte and est the min level at which the analyte can be quantified with acceptable accuracy and precision. A typical signal to noise ratio is 10:1
Validation of Analytical Procedures: Sensitivity
this is measured by deter the limit of detection (LOD). Can be done several ways such as: analyzing samples of known conc and est the min level at which the analyte can be reliably detected. OR using signal to noise ratios by comparing measured signals from samples with known low concentrations of analyte with those of blank samples and est the minimum conc at which the analyte can be detected, blanks must be analyzed to est background noise
Computer Tracking--Laboratory Information Management System (LIMS)
this is used to: track samples, test results, schedule work, store information about equipment, record standard operating procedures (SOPs)
Validation of Analytical Procedures: Specificity
this is when a test can discriminate bt compounds of closely related structures like drug metabolites. This can be demonstrated by obtaining positive results from the samples ctg the analyte and negative results from the samples that do not ctg the analyte. The assay should be applied to a like substance to check for specificity.
solid phase extraction systems
this is when the antibody to the drug is bound to the wall of a plastic tube, after incubation, the drug-antigen is bound to the wall of the tube, the bound drug can then be analyzed by gamma counting the tube or eluting the drug-antibody complex, re-suspending the soltn, and measuring any optical changes using an assay technique.
What is the seat of the fire?
this is where the fire started
Role of the Archivist
this person is responsible for the ID, maitenence, preservation, and use of all records. typically a member of the quality assurance team interested in both the physical preservation of documents and samples and also interested in the contexual relationship between all the items ex. GC-MS quantation of urinary drug levels that can be accessed and used in a forensic case
TLC : Stationary Phase
this phase is usually made of adsorbents (silica and alumina), drug separation is achieved through interactions bt the drug and the surface of stationary phase particles. Works through a partitioning process bt the stationary phase and the mobile phase. When stationary phase is more polar > mobile phase this is called a normal phase system When a mobile phase is more polar > stationary phase this is called a reverse-phase system
Establishing Accuracy
this should be established across the specified range of the analytical procedure. Can be achieved using several approaches: 1- best method is to use a reference std of known concentration 2-can also be inferred from precision, linearity, and specificity data--need a min of 9 determinations over a min of 3 conc levels Generally reported as the diff bt the mean and the accepted true value together with the confidence intervals Applies to the analytical method and the instrument
Internal Audit checklist
this type of audit can be a way to prepare for inspection by a regulatory agency, this can be done by lab personnel, one who is familiar with the equipment, but not specifically involved in that program. Typical checklist includes: names of staff members must haves--unique ID, COC, proper seal, samples protected, ID's based on 2 independant assays, amount of sample needed, security Receiving through confirmation: check new SOP's, interview employees, check freezers and fridge, check for samples left out or unattended, check receiving paper work, verify COC Check written policies---contamination issues, sample tampering, breakage, ect
in extraction recovery, before quantitative analysis can be performed, the recovery procedure is performed at a minimum of _____ concentrations
three
Fourier Transform Infrared Analysis (FTIR)
utilizes an interferometer instead of a monochromator, in this set-up the includes two mirrors where one is fixed and the other oscillates. This means that half of the light is reflected to the detector. The beams of light recombine and form an interferogram. When a sample is placed in the beam will absorb diff wavelengths of vibrational energy and the interferogram will be composed of all the wavelengths of light except those abs by the sample. A microscope is also placed at the lab bench so that analysts can obtain spectra from individual drug crystals. Sources--silicon carbide or nichrome coil Advantages--quick data collection, all frequencies detected simultaneously, increased sensitivity, increased spectral resolution, enhanced data manipulation
UV or Vis Components: Wavelength selector-monochromator
variable entrance slit, gives an optical image from the light source, a collimating lens or mirror that produces a paralell beam of radiation, a prism or refraction grating to disperse the radiation into it's component wavelengths, a focusing element the reform the image and an exit slit that isolates the spectral band
Interlaboratory transfer
when a sample is transferred to another laboratory facility it must accompanied by the original chain of custody info and any other info such as sample prep
Matrix-matched calibrations
when calibrations standards can be prepared in the appropriate matrix extract to provide a more accurate cailibration, but do not create the calibration curve with these, commonly used for pesticide residues, can be cost prohibative validity of the matrix used can be checked by adding a known qty of analyte to the sample extract and comparing the increase in detector response with the response produced by the matrix other issues that may arise--- 1. analyte is naturally present in all samples and only in levels higher than occur naturally are relevant ex. Br in celery 2. the analyte is of natural origin and detectable in most or all samples. ex. cabbage having CS2 3. the analyte is detectable in most, if not all samples but does not occur naturally ex. imazailil occurs post harvest in most citrus fruits 4. Background is due to an interfering natural or synthetic chemical aside from the analyte and is present in most samples. A more specific detection method should be used. Confirm residues with greater LCL (lowest calibration level) 5. In GC analysis matix-matching of calibration soltns and priming or conditioning are required
when must a lab conduct internal validation?
when developmental validation has been performed by another lab or at another site of a multi-lab system
Reference Standard Authenticity
when investigating an unexpected result, comparing to known results ex. comparison of known retention times or peaks at certain masses
Molar Absorptivity
when the conc is designated at mol/L
the three parts of the writing process that can yield useful information are
writing surface, writing instrument, and transfer medium
what is the definition of an SOP?
written document that details the method for an operation, analysis or action with thoroughly prescribed techniques and steps that is officially approved as the method for performing certain routine or repetitive tasks
in standard curve and y=mx+b, x = _________and y = _________
x = concentration, y = ratio of drug peak area to internal standard peak area
