Clin lab 1

Urine Sediment Examination

Casts - cylindrical appearance

Neurologic disorders

Multiple sclerosis Demyelinating disorders Guillain-Barré syndrome

Complete Metabolic Panel

A Comprehensive Metabolic Panel (CMP or "comp met") is another frequently ordered panel of chemistry tests that presents the provider with information on the patient's metabolism. The CMP includes all of the tests in the BMP plus albumin (ALB) alanine transaminase (ALT) alkaline phosphatase (ALP) aspartate aminotransferase (AST) bilirubin total protein (TP) Albumin comprises ~ 60% of the total protein within the extracellular portion of the blood (Hgb is the most abundant protein in whole blood and is intracellular) Transports many important blood constituents drugs, hormones, enzymes Albumin is synthesized in the liver and therefore is a measure of hepatic function An alanine aminotransferase (ALT) test measures the amount of this enzyme in the blood. This enzyme occurs mostly in hepatocytes with smaller quantities in skeletal and heart muscle. It is released into the circulation when cells are damaged or necrotic. Increased ALT usually indicates damage to the liver, although severe damage to skeletal muscle can produce significant elevations. Aspartate Aminotransferase (AST) is an enzyme that is present in hepatocytes and myocytes (both skeletal muscle and cardiac) Elevations in AST are most commonly a reflection of liver injury. It usually rises in concert with the ALT. When the AST is ≥2.0 times the ALT, alcohol abuse with cirrhosis or alcoholic hepatitis should be suspected. Alkaline phosphatase (ALP) includes a number of cellular enzymes that hydrolyze phosphate esters. It is an enzyme present in a number of tissues, including liver, bone, kidney, intestine, and placenta, each of which contains distinct isoenzyme forms Isoenzymes are forms of an enzyme that catalyze the same reaction, but are slightly different in structure The two major circulating alkaline phosphatase isoenzymes are bone and liver. Therefore elevation in serum alkaline phosphatase is most commonly a reflection of liver or bone disorders. Waste product from breakdown of heme in the liver Heme is component of hemoglobin Total serum bilirubin level is the sum of the conjugated (direct) and unconjugated (indirect) bilirubin. Normally the unconjugated bilirubin makes up 70-85% of the total bilirubin Used to evaluate liver function or diagnose anemias caused by red cell destruction Total protein measures 2 classes of protein— albumin & globulin Most serum proteins are synthesized in the liver (albumin and many others) or by mature plasma cells (immunoglobulins). Increases or decreases in serum proteins represent a balance between synthesis and protein catabolism

Semen Analysis

A semen analysis focuses on three main areas - motility, morphology, sperm count - to determine: Infertility Successful vasectomy Successful vasectomy reversal

Urinalysis Panel

A urinalysis screen is a group of 10 chemical tests used to detect urinary tract infections, as well as metabolic and kidney disorders.

Unpredictable Dose Response

An unpredictable dose response occurs when the same dose produces a therapeutic effect in one patient and toxic effects in another, or when a dose adjustment in the same patient produces a response beyond what was anticipated

encephalitis

Bacterial Listeria, Leptospira, Borrelia, Treponema Viral - Most common cause Herpesviridae, Enteroviruses , Arboviruses Rabies Fungal Histoplasma, Cryptococcus Parasitic Toxoplasma, Taenia

Meningitis

Bacterial: Life threatening; vaccine preventable Viral Self-limiting Fungal AIDS defining illness Parasitic Free living amebae 99.999% fatal Mycobacterial

CSF test

Chemistry tests Protein ↑ levels seen with meningitis, hemorrhage, and multiple sclerosis ↓ levels associated with leakage of fluid out of the CNS Glucose Tested immediately ↓ seen with bacterial meningitis, but not viral Glutamine produced from ammonia and a-ketoglutarate by the brain cells. ↑ Liver disorders, in directly detects excess ammonia within CNS Lactate ↑ Seen in bacterial, fungal, and tubercular meningitis In bacterial, tubercular, and fungal meningitis, CSF lactate levels greater than 25 mg/dL occurs much more consistently than does decreased glucose. Levels greater than 35 mg/dL are frequently seen with bacterial meningitis, whereas in viral meningitis, lactate levels remain lower than 25 mg/dL Microbiological examination CSF is normally sterile Gram Stain (<1 hr) Culture (72 hours) Antigen tests Bacterial antigen Cryptococcal antigen Serology testing - useful when PCR is negative Typically for neurosyphilis Can ONLY use the VDRL test for CSF workup Venereal Disease Research Laboratory - non-treponemal test similar to RPR Other pathogens Measles virus Rubella virus West Nile virus Toxoplasma gondii Herpes Simplex virus Varicella Zoster virus

Cut points

Choosing the cut off - Choice depends on relative implications of false positive and false negative errors Choose a low cut-point (= increase sensitivity) If results of missing a case (FN) are important If cost of further diagnostic confirmation is not high e.g. phenylketonuria PKU test. Choose a higher cut-point (= increase specificity) If implications of a false positive are serious e.g., alpha-fetoprotein test for Down's syndrome during pregnancy

Common Wet Mount Microscopic Findings

Clue Cells: Clue cells are epithelial cells of the vaginal wall. They often appear fuzzy, lacking distinct sharp borders due to being covered with bacteria. Clue cells are a sign of bacterial vaginosis caused by high concentrations of Gardnerella vaginalis, Mycoplasma hominis and anaerobic bacteria such as Prevotella sp. and Mobiluncus sp. In the images below, the arrows indicate examples of clue cells.

Storage Containers

Containers/conditions 24 hr urines (amber colored plastic bottle + preservatives) Blood parasites (e.g. malaria) - EDTA lavender top tube Acute viral hepatitis panel - serum separator tube (SST) Transport containers Stool culture (> 2 hrs) - Cary Blair Transport media Chlamydia/Gonorrhea - manufacturer urine collection tube

Ways of Assessing Validity

Content or "Face" validity: does it make clinical or biological sense? Does it include the relevant symptoms? Criterion: comparison to a "gold standard" definitive measure (e.g., biopsy, autopsy) Expressed as sensitivity and specificity Construct validity (this is used with abstract themes, such as "quality of life" for which there is no definitive standard) Diagnostic judgments are based on probabilities That using a quantitative approach is better than just guessing! That you will gradually become familiar with the typical accuracy of measurements in your chosen clinical field; That the principles apply to both diagnostic and screening tests; Of some of the ways to describe the accuracy of a measurement. Reliability: consistency or reproducibility; this considers chance or random errors (which sometimes increase, sometimes decrease, scores). "Is it measuring something?" Validity: "Is it measuring what it is supposed to measure?" By extension, "what diagnostic conclusion can I draw from a particular score on this test?" Validity may be affected by bias, which refers to systematic errors (these fall in a certain direction) Safety, Acceptability, Cost, etc.

Potassium hydroxide (KOH Prep)

Dissolves tissue material (debris and cells) in vaginal secretions, nails, and skin scrapings, only fungal elements remain intact Often performed after the saline prep

Waived Tests

Employ methodologies that are so simple and accurate as to render the likelihood of erroneous results negligible Pose no reasonable risk of harm to the patient if the test is performed incorrectly; Are cleared by the Food and Drug Administration for home use Dipstick or tablet reagent urinalysis Fecal occult blood Ovulation tests Urine pregnancy tests Erythrocyte sedimentation rate Hemoglobin Most "bed side" testing

Analytical Sources of Error

Error rate is close to 3.2 sigma performance (95%) Quality controls not prepared correctly Calibrations performed incorrectly Maintenance not performed on instruments Controls and calibrations interpreted incorrectly Wrong test performed on specimen Testing personnel misinterpretation of results

Polarizing Light Microscope

Facilitates the study of unstained structures, which can still be alive. Similar to a bright-field microscope, except has changes to the condenser and objective.

Serous Fluids (Pleural, Pericardial, and Peritoneal)

Fill the space in the closed cavities of the body to lubricate the membrane surfaces Normally pale and yellow ultra-filtrates of plasma ↑ in volume of these fluids is an effusion Serous fluid is collected by needle aspiration from the respective cavity: Thoracentesis: pleural cavity Pericardiocentesis: pericardial cavity Paracentesis: peritoneal cavity Two types of effusions TRANSUDATE = Effusions that form because of a systemic disorder that disrupts the balance in the regulation of fluid filtration and re-absorption Example: changes in hydrostatic pressure created by congestive heart failure or the hypoproteinemia associated with the nephrotic syndrome. EXUDATE = Exudates are produced by conditions that directly involve the membranes of the particular cavity LOCAL accumulation of fluid due to increased vascular permeability due to cells caused by blocked blood vessels or lymph vessels, inflammation, tissue injury, and tumors

Amniotic Fluid

Fluid found in membranous sac that surrounds the fetus Provides a cushion to protect the fetus Enables fetal movement Plays a role in biochemical processes Amniocentesis: collection of fluid by needle aspiration from the amniotic sac PPM - Fern test can identify ruptured membranes Not the same as amniocentesis Chromosomal Defects Cytogenetic evaluation will detect chromosomal abnormalities and provide the gender of fetus Cells in the fluid are cultured and chromosome analysis (karyotype) is performed Neural Tube Defects Alpha-fetoprotein (AFP) level can be measured to detect possible neural tube defects (spina bifida and anencephaly). AFP is a substance made in the liver of the fetus. May also be tested from maternal blood or urine. Hemolytic Disease Evaluation of the bilirubin level is used to detect abnormal fetal RBC hemolysis due to Hemolytic Disease of the Newborn (HDN) Fetal Pulmonary Development Measurement of fetal alveolar surfactant is used to evaluate fetal lung maturity

Clinical Pathology

Hematology - the study of blood and its disorders , looks specifically at blood components such as cell counts, and blood and bone marrow cells Aids to diagnose anemia, hemophilia, blood-clotting disorders, and leukemia Urinalysis - Performs routine urine screening tests for the detection of disease related to the kidneys and urinary tract Chemistry - uses chemical processes to measure levels of chemical components in the blood most common specimens are blood and urine Tests include blood glucose, electrolytes, enzymes, hormones, lipids other metabolic substances, and proteins Transfusion Services - ensures donated blood or blood products are safe, before they are used in blood transfusions includes typing the blood for transfusion and testing for infectious diseases Microbiology - is the study of disease-causing microorganisms (bacteria, fungi, parasites, viruses) is responsible for identifying infectious agents in blood, urine, sputum, feces, cerebrospinal fluid and other body fluids uses various techniques to identify microorganisms, including: culture chemical, immunological, and genetic tests examination under a microscope staining Immunology and Serology - Immunology is the study of the body's immune system and its functions and disorders Serology is the study of blood serum Immunology and serology tests focus on: identifying antibodies investigating problems with the immune system determining organ compatibility

Other Laboratory Panels

Hepatic function panel Thyroid panel Obstetric triple/quad screen Dementia screening Sexually transmitted diseases panel Myocardial infarction panel Renal function panel Drugs of abuse screen Bleeding disorders panel

Screening testing

Hight specificity

Rejection Criteria

Improperly collected and improperly labeled. Prolonged transport - degraded Leakage Incomplete requests Clotted, hemolysis, or quantity not sufficient. Non-fasting specimen Improperly preserved specimen Specimens unprotected from light Acidified specimen Must be frozen.

Quality Assessment-Proficiency testing

Incorporated into the CLIA requirements Periodically the laboratory will conduct testing of samples sent from "outside" organizations Sample values are unknown to the testing laboratory Samples must be tested and results reported as if they were patient samples No cheating allowed!!! Results by committee; Getting help from other labs; Sending specimen off for someone else to perform test The testing laboratory is graded on its performance for testing the unknown samples A laboratory that incorrectly reports results, e.g., "fails" the proficiency test, may loose its accreditation to perform those tests Not meeting the standard = Shut down testing for that specific analyte $$$ ramifications, lack of confidence by staff

Characteristics of an Ideal Drug for Therapeutic Drug Monitoring

Readily available & accurate assay exists Strong correlation exists between the drug concentration and efficacy or toxicity Lacks easily observable, safe, or desired clinical endpoints (e.g., seizure, arrhythmia) Possesses a narrow therapeutic index or range Demonstrates an unpredictable dose-response relationship (e.g., non-linear pharmacokinetics)

Meningitis and Encephalitis

Inflammation of the meninges = meningitis sudden fever, severe headache, nausea/vomiting, double vision, drowsiness, sensitivity to bright light, and a stiff neck Inflammation of the brain = encephalitis fever, seizures, change in behavior, confusion and disorientation The majority of infectious encephalitis cases are caused by viral infections

Distribution

Insoluble drugs require a protein carrier Two most common carrier proteins Albumin Alpha1-Acid Glycoprotein Both proteins are produced by the liver Presence of any liver disease will increase the free fraction of the drug(s) Free fraction The therapeutically (biologically) active fraction of the drug (non-protein bound) Bound and free fractions exist in equilibrium Most laboratory assays measure the total drug concentration (the sum of free and bound)

Quality control statistics

Mean: Often referred to as the "average" Median: Represents the middle values in your list 2, 4, 6, 8, 10 Mode: The mode in a list of numbers refers to the list of numbers that occur most frequently 5,2,6,2,3,3,2 Standard deviation: measures the amount of variation from the average within 2 standard deviations Each laboratory establishes their own reference ranges Pathological range diagnoses disease AKA reference range (based on a bell curve) Functional range assesses risk for disease before the disease develops. Your labs may be "normal" but you still feel sick

Monosodium Urate Crystals (Gout)

Monosodium urate crystals are seen in gout/gouty arthritis and appear as needle-shaped negatively birefringent crystals varying in length from 2 to 20 microns. With negative birefringence, the crystals appear yellow in parallel light and blue with perpendicular light.

Nasal Smears

Nasal smear eosinophilia contributes to the diagnosis of allergic rhinitis

Interpreting Results Drug test

Negative Test Drug free Drug present but below screening cutoff level Values below cutoff are not reported Positive Test Positive initial screening result Positive rescreening result Positive GC/MS result Intact chain of custody

Storage Temperature

Optimal conditions for sample viability Glucose is metabolized at room temperature at a rate of 7 mg/dL/hour (0.4 mmol/L/hour); at 4° C, the loss is approximately 2 mg/dL/hour. Refrigeration Urine (>2 hrs) Room Temperature Cerebrospinal fluid CSF Frozen Plasma ammonia submit on ice 15 minutes

Testing CSF

Puncture at the interspace of vertebrae L3 and L4 or lower - always STAT test Open pressure reading - The patient must be in the lateral recumbent position. Attach the manometer through the stopcock, and note the height of the fluid column. Normal opening pressure is 5-20 cm water Collecting CSF - Collect at least 10 drops of CSF in each of the 4 plastic tubes - up to 20 ml total If the volume of fluid obtained is limited, tests may need to be prioritized. Collection of cerebrospinal fluid Tube 1: chemical and serologic tests Glucose, protein, etc Tube 2: Microbiology Gram stain, cultures Tube 3: Hematology Total cell counts and differential count Tube 4: A fourth tube may be drawn for the microbiology laboratory to better exclude skin contamination or for additional serologic tests. Hematology tubes are refrigerated. Microbiology tubes remain at room temperature. Chemistry and serology tubes are frozen. Routine examination of cerebrospinal fluid Gross appearance Red and white blood cell counts Morphologic examination Chemistry tests Microbiological examination Serology tests Gross appearance Turbidity (WBCs, bacteria, protein, lipid) Clots (Protein, traumatic tap) Color (successive tubes should show less color) Xanthochromia: pale pink, orange or yellow color in the supernatant Red and white blood cell counts Normally no RBCs in CSF SUBARACHNOID BLEED subarachnoid hemorrhage Elevated opening pressure Presence of RBCs Presence of xanthochromia WBCs normal at 0-8 cells/µL Morphologic examination Cytocentrifugation Smears from centrifuged spinal fluid sediment Differential cell count

Serous Fluids examination

Routine examination of serous fluids Gross appearance Clotting? Typically indicates high protein level Red and white blood cell counts Morphologic examination and white cell differential Microbiological examination Chemical analysis Protein Lactate dehydrogenase Glucose Visual appearance Normal - clear and straw colored with low viscosity Abnormal - turbid, viscous, colored Turbidity - high WBC count Turbid > 200 cells/ul, Purulent 10,000 cells/ul Viscous - clotting and/or hydrolyzed debris Red or pink color - presence of blood No history of trauma suggests malignancy Cytologic examination for PMS, Lymphocytes, Monocytes, malignant cells Culture of microorganisms when infection is suspected (Gram stain, culture, AFB smear) Protein concentration estimated from specific gravity measurement Normal or transudate: s.g. < 1.015 and protein concentration < 2.0 g/dl Exudate: s.g. > 1.018 and protein concentration > 3.0 g/dl Protein concentration Normal in transudates Increased in exudates Glucose estimated from relationship between protein concentration and leukocyte count Normal protein <=> low leukocyte count <=> normal glucose Elevated protein <=> high leukocyte count <=> decreased glucose Glucose concentration Normal in transudates Decreased in exudates LDH level is increased in exudates from damaged tissue and dead leukocytes. LDH level is normal in transudates, except when a lymph duct drains a tumor; the increased LDH results from liberation from rapidly turning over neoplastic tissue.

Seminal fluid (semen)

Semen analysis is primarily used to Evaluate post-vasectomy patients to insure post-operative infertility Evaluate reverse-vasectomy patients Evaluate primary infertility as part of the work-up of both the male and female Detailed instructions provided to the patient by the laboratory Following 3 days of sexual abstinence, an entire ejaculate is collected in a sterile container The collection should be done near a laboratory and delivered immediately after collection Specimens collected elsewhere should be kept at room temperature and delivered to lab no more than 30 minutes following collection A delay in testing will decrease sperm motility The exact time of collection must be recorded as semen liquefaction is one of the parameters tested Post vasectomy - Evaluated for the presence or absence of sperm If sperm are present, sperm viability is recorded as "motile" or "non-motile" Infertility work up - Color: grayish-white and opalescent Volume: 2-5 ml Viscosity/Liquefaction: initially viscous, but will become watery within 60 minutes of collection

POC Testing

Shorten turnaround time for critical results and enhance convenience for both patients and caregivers. Usually 30 min or less. CLIA waived tests are typically exempt from most regulations involving personnel, proficiency testing, and rigorous quality assurance requirements. Laboratory oversight is mandatory Method validation, training, policies and procedures, responsibilities. Limitations - Technology - accuracy, precision, reliability Knowledge and training Quality assurance Accountability - ordering, reporting, data management Cost - capital, consumables, staff Connectivity

Specimen Collection Tool

Specimen Collection Guidelines Lab Submission Guide Types of Specimens Volume Timing of Collection Storage Specimen Rejection Criteria Transport Proprietary requirements Add-on tests Reflex tests Legal Issues

Brain Hemorrhage

Subarachnoid (SAH) Bleeding in the subarachnoid space Aneurysms and head trauma Intra-cerebral Rupture of small vessels within the brain Chronic high blood pressure Cerebral infarct Blockage of blood flow to brain

Fern Test

The Fern Test is used to detect amniotic fluid leakage from the membrane surrounding the fetus during pregnancy. If rupture of the membrane has occurred evidence of amniotic fluid will be present. Normal vaginal secretions that do not contain amniotic fluid will not show the ferning pattern. The presence of amniotic fluid "fern-like" pattern (crystallization) on a slide viewed under low power on a microscope

Unacceptable Specimens

The big three: Icteric (jaundice) specimens Lipemic specimens Hemolyzed specimens

Calcium Pyrophosphate Crystals(Pseudogout)

The crystals appear blue in parallel light and yellow with perpendicular light. Opposite of monsodium urate crystals.

Basic Metabolic Panel (BMP)

The BMP is a chemistry panel. The BMP includes electrolytes and tests of kidney function: Sodium (Na+) - Together with Cl−, it makes the major contribution to the plasma osmotic pressure. Increased serum sodium level = Hypernatremia: This always indicates a relative total body water deficit, regardless of the extracellular volume status Decreased serum sodium level = Hyponatremia: This always indicates a relative total body water excess, from excess water ingestion or inability of the kidney to excrete a sufficiently dilute urine, regardless of the extracellular volume status Potassium (K+) - Helps maintain the body's balance of fluids; also important in muscle & nerve function Plasma [K+] is tightly regulated by the kidney: hyperkalemia leads to aldosterone secretion and potassium excretion; hypokalemia leads to excretion of urine nearly devoid of potassium Elevated serum potassium level = Hyperkalemia: Primary and secondary hypoaldosteronism Decreased serum potassium level = Hypokalemia: Diabetes mellitus, Cushing syndrome, hyperaldosteronism Chloride (Cl-) - Chloride is the major extracellular anion with serum concentration of ~ 100 mmol/L Important electrolyte that helps maintain normal functions and the body's balance of fluids Hyperchloremia and hypochloremia are rarely isolated phenomena. Usually they are part of shifts in sodium or bicarbonate to maintain electrical neutrality. Carbon Dioxide Content (CO2) - CO2 + H2O ↔ H2CO3 ↔ H+ + HCO3- The carbon dioxide content (CO2) measures the H2CO3, dissolved CO2 and bicarbonate ion (HCO3) that exists in the serum. Because the amounts of H2CO3 and dissolved CO2 in the serum are so small, the CO2 content is an indirect measure of the HCO3 anion Increased bicarbonate This indicates a metabolic alkalosis, either primary or secondary to a respiratory acidosis Decreased bicarbonate A decreased bicarbonate concentration indicates the presence of a metabolic acidosis. Blood Urea Nitrogen (BUN) - The BUN measures the amount of urea nitrogen in the blood. Urea is formed in the liver as the end product of protein metabolism and is transported to the kidneys for excretion. An increase indicates decreased glomerular filtration and/or increased tubular reabsorption, inadequate excretion or increased production in the gut from ingested protein or blood. The BUN is interpreted in conjunction with the creatinine test . These tests are referred to as "renal function studies". Serum Creatinine (Cr) - The creatinine test measures the amount of creatinine in the blood. Creatinine is a catabolic product of creatine phosphate used in skeletal muscle contraction. Creatinine, as with blood urea nitrogen, is excreted entirely by the kidneys and blood levels are therefore proportional to renal excretory function. Serum glucose (Glu) - Plasma glucose levels should be evaluated in relation to a patient's meal i.e., postprandial vs fasting Elevated glucose levels may also be indicative of diabetes mellitus Glucose is one of the most commonly measured test in the laboratory Total Calcium (Calcium) - The total serum calcium is a measure of both Free (ionized) calcium Protein bound (usually to albumin) calcium Therefore, the total serum calcium level is affected by changes in serum albumin As a rule of thumb, the total serum calcium level decreases by approximately 0.8mg for every 1 gram decrease in the serum albumin level. The plasma level varies with the rate of Ca2+ absorption from the small intestine and the proximal renal tubular reabsorption rate, under the control of parathyroid hormone (PTH).

Venous Blood Samples

The most common specimen submitted to the laboratory (approximately 80% of the laboratory workload) Second most common is capillary (skin puncture) Composition is a mixture of arteriole, capillary, and venule blood in addition to tissue fluid(interstitial/intracellular) Performed when venipuncture is contraindicated. -eg. Babies, small children, burn victims

Pre-Analytical

The most important component of laboratory medicine. Inclusive of all the steps that must take place before a sample can be analyzed. Account for up to 70 % of all lab errors. Effects Increased overall costs - recollection, retesting, unnecessary workups Strict adherence to proper guidelines can minimize collection variables. Sample misidentification Improper timing Improper fasting Wrong anticoagulant to blood ratio, Improper mixing Incorrect order of draw Hemolyzed or lipemic specimens Short fill sample tube Wrong specimen container or preservatives

Postcoital Test (PCT)

The post coital test (PCT) is a test in the evaluation of infertility Cervical mucus is collected 2 to 8 hours after coitus and evaluated for consistency of cervical mucus and ferning as well as the ability of spermatozoa to penetrate the mucus and maintain activity

Therapeutic Range

The therapeutic range is the range of drug concentrations within which the probability of the desired clinical response is high and the probability of an unacceptable toxicity is relative low. Minimum Effective Concentration Below this level, therapeutic effect is not achieved Minimum Toxic Concentration Above this level, symptoms of toxicity appear

Trough And Peak Values

Trough value Lowest therapeutic concentration achieved during a dosing cycle Typically occurs shortly before the next dose and should be above the Minimum Effective Concentration of the drug Peak value - toxicity risk Highest therapeutic concentration achieved during a dosing cycle Should be below the Minimum Toxic Concentration of the drug

Fluorescence Microscope

Ultraviolet illumination Visualization of specimens that contain naturally fluorescent substances or that have been stained with fluorescent stains/dyes Used in diagnosis of infectious diseases Applied to identify specific antibodies in response to antigens Similar to a darkfield microscope with the addition of wavelength selection. Objects that fluoresce absorb light at short (ultraviolet wavelengths), but emit light at longer (visible) wavelengths. Typically used with florescent antibody techniques Autoantibodies directed against contents of the cell nucleus. They are present in higher than normal numbers in autoimmune disease. The ANA test measures the pattern and amount of autoantibody

Synovial Fluid

Viscous fluid found in joint cavities which is formed as an ultrafiltrate of plasma across the synovial membrane The presence of the mucopolysaccharide hyaluronic acid differentiates synovial fluid from other fluids in the body Forms a thin layer 50 μm at the surface of cartilage Reduction of friction - synovial fluid lubricates the articulating joints Shock absorption - as a dilatant fluid Viscosity increases with the rate of shear strain Nutrient and waste transportation - the fluid supplies oxygen and nutrients and removes carbon dioxide and metabolic wastes Arthrocentesis Collection of synovial fluid by needle aspiration Classifications of synovial fluid - Class I Noninflammatory - degenerative diseases like osteoarthritis Class II Inflammatory - associated with immunologic diseases like lupus and rheumatoid arthritis(ELEVATED PROTIENS THAT ARE ANTIBODIES) Class III Infectious - bacterial infection Hemorrhagic - traumatic injury, tumor, or coagulation deficiencies (hemophilia). Crystal-induced - gout and pseudogout Routine examination of synovial fluid Gross appearance Color and clarity Viscosity Red and white blood cell counts Morphologic examination Microscopic examination for crystals Brightfield or phase-contrast microscopic examination Polarized light microscopy Microbiological examination All synovial fluids should have a Gram stain and culture performed because acute bacterial arthritis is the most rapidly destructive disease of joints Chemistry tests Glucose Protein Other tests Immunologic tests Typical for suspected rheumatoid arthritis or SLE Antinuclear antibodies (ANA)

Post-Analytical

What to do with the result This is usually where we forget how to communicate with each other Critical Test value - must be reported within 1 hr of completion (telephonically) to ordering provider STAT test result - must be reported within 1 hour of RECEIPT (usually telephone if critical)

CBC + Differential

When a differential is ordered, the percentage of each type of leukocyte present in a specimen is measured. Types of leukocytes Neutrophils (PMNs) Lymphocytes Monocytes Eosinophils Basophils WBC differentials are either performed manually or by an automated instrument

Common Wet Mount Microscopic Findings

White Blood Cells White blood cells (WBCs) are a normal component of vaginal flora. WBCs are small and have a multi-lobed nucleus and appear dark and granular. WBCs can be elevated in infections involving Chlamydia, Trichomonas vaginitis, herpes, and Neisseria gonorrhoeae. A ratio of one WBC for every epithelial cell is considered within normal limits. In the images below, the arrows indicate examples of white blood cells.

Types of Blood Specimens

Whole Blood Formed elements and fluid plasma remain combined (sample does not clot) Serum Blood clots (w/in 20-30 min) consisting of blood cells trapped in a mesh of fibrin - leaving serum The liquid portion of clotted blood samples Lacks coagulation protein fibrinogen Plasma Blood must NOT clot (anticoagulant added) Centrifugation forces RBCs down and leaves plasma on top The liquid portion of anticoagulated whole blood Contains all coagulation proteins

Common Wet Mount Microscopic Findings

Yeast: Yeast are unicellular fungi that appear commonly in vaginal discharge wet mounts. Yeast can appear as budding yeast or yeast with pseudohyphae, an elongated filament-like string of attached cells. In the images below, the arrows indicate examples of yeast.

Common Laboratory Panels

Basic Metabolic Panel (BMP) Comprehensive Metabolic Panel (CMP) Complete Blood Count (CBC) Urinalysis Lipid

Dark-field Microscopes

Darkfield describes an illumination technique used to enhance the contrast in unstained samples. It works by illuminating the sample with light that will not be collected by the objective lens, and thus will not form part of the image. This produces the classic appearance of a dark, almost black, background with bright objects on it Light passes across the specimen resulting in no direct light coming through the objective. Typical for viewing spirochetes Used to view unfixed, unstained specimens Such as living organisms - Syphilis Background dark, specimen bright

Saline Wet Prep

Direct wet mounts for the presence or absence of bacteria, fungi, parasites, and human cellular elements Commonly used for vaginal smears to look for the presence of Trichomonas vaginalis, clue cells, and yeast (causative agents of vaginitis) Trichomonas vaginalis:Trichomoniasis is a sexually transmitted infection caused by the parasite Trichomonas vaginalis, a motile pear-shaped protozoan. Trichomoniasis causes a vaginal discharge that is yellow-green, foamy, with an odor. In the images below, the arrows indicate examples of the Trichomonas vaginalis parasite.

Fecal Leukocytes

Fecal leukocytosis is a response to infection with microorganisms that invade tissue or produce toxins, which causes tissue damage and is a good indicator of the presence of an invasive microbiological pathogen such as Salmonella or Shigella.

Quality Assurance/Assessment (QA)

Follow procedures

1988 Clinical Laboratory Improvement Amendments (CLIA)

Four categories of test complexity as determined by the Food and Drug Administration (FDA): Minimal Complexity (commonly referred to as waived testing) Provider Performed Microscopy (a subcategory of moderate complexity testing) Moderate Complexity High Complexity

Functions/Properties of the BBB

Functions Protects against "foreign substances" Protects against hormones and neurotransmitters Maintains a constant environment Properties Large molecules do not pass BBB Lipid insoluble molecules do not cross, but lipid soluble molecules will High electrical charged molecules are slowed Testing indications - Meningeal infection Brain hemorrhage Primary or metastatic malignancy Neurological diseases

Toxicology Specimens

Gastric contents Best when the time interval between ingestion of the toxin and presentation to a medical facility is short Blood Best for detection of acetaminophen, ethanol, salicylates, carbon monoxide (carboxyhemoglobin levels), pesticides, and lead Urine Best for detection of arsenic, mercury, and drugs of abuse

Drug Metabolism and Elimination

Metabolism typically renders non-polar lipophilic drugs into more polar (water soluble) compounds for elimination Primary site for drug metabolism → Liver Polar, non-lipophilic drugs are eliminated primarily through renal excretion, which is dependent on adequate renal function, renal blood flow, urine pH, and drug molecule size Less common elimination pathways → feces, sweat, and respiration Drug clearance is the theoretic volume of plasma that is completely cleared of a drug per unit of time Clearance is the sum of all elimination mechanisms: Hepatic Renal Biliary And any other specific to that drug Patients with impaired clearance need more monitoring!

Provider Performed Microscopy (PPM)

Moderate complexity Menu: Urine sediment examinations Direct wet mounts for the presence or absence of bacteria, fungi, parasites, and human cellular elements Potassium hydroxide prep (KOH) - fungal elements Pinworm examinations Fern tests Postcoital direct qualitative exam of vaginal or cervical mucus Nasal smears for granulocytes Fecal leukocyte examinations Qualitative semen analysis Must be personally performed by the provider Must be categorized as moderately complex Primary instrument is the microscope The specimen is labile Controls are not available There is limited specimen handling

Common routes

Oral The GI tract must have the ability to absorb drugs that are administered orally The ideal drug must withstand the acidity of gastric fluids and be readily transported across the lipid membranes of the intestinal wall Suppository Commonly used in infants when an oral form of a drug (usually a liquid) is not available Parenteral The best route for drugs unable to survive exposure to gastric fluids or first-pass elimination Intravenous: injected into a vein Intramuscular: injected into muscle Subcutaneous: injected under the skin Transdermal Drug is applied to the skin and absorbed systemically

Order of draw

Pediatric draw we reverse order - purple , green , red Green in ran quicker

Additional terms

Positive predictive value (PPV) The proportion of subjects with positive test results who are correctly diagnosed Negative predictive value (NPV) The proportion of subjects with a negative test result who are correctly diagnosed Prevalence: Apparent verus True Apparent: The total number of positive(TP+FP) cases in the population, divided by the number of individuals in the population True: actual number of diseased individuals(TP+FN) divided by the number of individuals in the population Higher prevalence increases PPV Lower prevalence increases NPV Prevalence does NOT affect sensitivity or specificity Positive Predictive Value (PPV) = TP/(TP+FP) = Probability that a positive score is a true positive NPV = TN/(TN+FN); same for a negative test result BUT... there's a big catch: We are now working across the columns, so PPV & NPV depend on how many cases of disease there are (prevalence). As prevalence goes down, PPV goes down (it's harder to find the smaller number of cases) and NPV rises. So, PPV and NPV must be determined for each clinical setting,

TDM Specimen Requirements

Specimen of choice = serum (Red top) Specimens must be drawn at the appropriate time Peak specimens are usually drawn shortly after a dose is given (medication-specific) Trough specimens are drawn shortly before the next dose

Common Wet Mount Microscopic Findings

Squamous Epithelial Cells: The majority of cells observed in a normal vaginal wet prep will be vaginal epithelial cells. Squamous epithelial cells are large and flat with an irregular shape, distinct borders, and a single nucleus. In the images below, the arrows indicate examples of squamous epithelial cells.

Laboratory Departments/Divisions/Sections

Support Services - Central Process/Client Services Phlebotomy Clerical Services Laboratory Information Systems

Toxicology

The study of poisons, their actions, their detection and the treatment of the conditions produced by them ~50%: suicide attempts ~30%: accidental (mainly children) ~20%: occupational exposure and homicide Common drugs and toxins tested Acetaminophen Carboxyhemoglobin (CO) Ethanol Salicylates Drugs of abuse Lead Selection of the best specimen is based on: When and how the patient was exposed The mode of exposure (injected, ingested) In general, specimens for ingested toxins are time-dependent Short time since ingestion - gastric Moderate time since ingestion - blood Long time since ingestion - urine It makes no sense to test a gastric specimen for the presence of an injected toxin

Post Analytical Add-On Tests

"Additional tests" requested after a specimen has been previously collected. Problems Quantity insufficient 3 mL of a specimen, (2nd or 3rd test total combined volume required is > 3 mL) Degradation of the analyte Glucose degradation (storage temp slide) Different anticoagulant or additive Red top specimen originally collected, new test requires purple top specimen Screening and confirmatory tests HIV ELISA + Western blot Syphilis RPR + MHA-TP Hepatitis Acute virus hepatitis panel (positive result) + Identifying hepatitis test Urinalysis Dipstick + Culture

Recommendations why one should order tests

- Identify changes in a patient's health - Diagnose a disease or condition - Plan treatment for a disease or condition - Evaluate the response to a treatment - Monitor the course of a disease over time interpreted within the context of a patient's overall health

Common Wet Mount Microscopic Findings

Bacteria: Bacterial vaginitis is often associated with a thin, gray-white or milky vaginal discharge adhering to the vaginal wall to give a fishy smell. Bacterial vaginitis is caused by high concentrations of Gardnerella vaginalis, Mycoplasma hominis and anaerobic bacteria such as Prevotella sp. and Mobiluncus sp. In the images below, the arrows indicate examples of bacteria.

Body Fluid Analysis

Body fluids in this lecture: Cerebrospinal Fluid (CSF) Synovial Fluid Amniotic Fluid Seminal Fluid Serous Fluid Pleural Fluid, Pericardial Fluid, Peritoneal Fluid

Light Microscopes

Bright-field microscope Background is lighter than observed specimen Most specimens require fixing and staining for bright-field microscopy Use visible light and optical lenses Either simple or compound Ocular lens Objective lens Final magnification: Multiply the enlarging power of both the ocular and objective lenses The image seen with this type of microscope is two dimensional. This microscope is the most commonly used in laboratory setting. It has high magnification. However, it has a low resolution. It is necessary to stain or dye most samples to give sufficient contrast and detail.

Urine Sediment Examination

Cells - RBCs, WBCs, epithelial cells RBCs - increased numbers indicate a variety of urinary tract and systemic conditions WBCs - pyuria, indicative of infection Epithelial cells - Squamous type - usually insignificant; Renal Epithelial -significant Casts: Urinary casts are small cylindrical structures that can be found in urine. Casts are formed by the solidification of proteins in the lumen of the kidney tubules and vary in size and shape according to the tubules where they were formed. The presence of casts in urine is associated with various pathologic conditions such as glomerular or tubular damage, renal inflammation or infection.

Evacuated Blood Collection Tubes

Color coded caps identify which additives and/or anticoagulants are present Vacuum will draw in a set volume of blood Invert tubes for sufficient contact with the additive - inaccurate results Order of draw must be followed to eliminate the possibility of cross contamination

Urine Sediment Examination

Crystals - form by precipitation of urinary salts, often due to pH, temperature, concentration Crystals in normal acid urine Uric acid, Calcium oxalate Crystals in normal alkaline urine Calcium phosphate, Magnesium phosphate, Calcium carbonate, Ammonium biurate Crystals in abnormal urine Cystine, Tyrosine, Leucine, Sulfonamide, Ampicillin

Pre-Collection Variables - Physiology

Diurnal variation - AM vs PM Testing for hormones, iron, acid phosphatase, and urinary excretion of most electrolytes such as sodium, potassium, and phosphate Cortisol - Peaks 4-6 AM; lowest 8 PM-12 AM; 50% lower at 8 PM than at 8 AM; increased with stress Insulin - lower at night Iron - Peaks early to late morning; decreases up to 30% during the day Exercise Physical activity has transient and long-term effects on laboratory determinations Exercise may elevate creatine phosphokinase (CK), aspartate aminotransferase (AST), and lactate dehydrogenase (LD), and may activate coagulation, fibrinolysis, and platelets These changes are related to increased metabolic activities for energy purposes and usually return to pre-exercise levels soon after exercise cessation. Diet Glucose and triglycerides, absorbed from food, ↑ after eating - Fasting Blood Glucose Long-time vegetarian diet ↓ lipid panel results High meat or protein-rich diet ↑ serum urea, ammonia, and urate levels High protein, low carbohydrate diet ↑ ketones in the urine and serum blood urea nitrogen (BUN). Ethanol ingestion ↑ plasma lactate, urate, and triglyceride concentrations Stress Mental and physical stresses induce the production of adrenocorticotropic hormone (ACTH), cortisol, and catecholamines. Mild stress induces an increase in total cholesterol while decreasing HDL cholesterol Hyperventilation affects acid-base balance and elevates leukocyte counts, serum lactate, or free fatty acids. Posture Elements that are affected by postural changes are albumin, total protein, enzymes, calcium, bilirubin, cholesterol, triglycerides, and drugs bound to proteins. An upright position ↑ hydrostatic pressure, which will ↓ plasma volume and ↑ concentration of proteins. Albumin and calcium levels ↑ when changing position from supine to upright. Bed rest can ↓ patient's hemoglobin (Hb) compared to the original admitting value enough to falsely lead a physician to suspect internal hemorrhage or hemolysis Age Bilirubin concentration rises after birth and peaks at about 5 days. In cases of hemolytic disease of the fetus and newborn (HDFN), bilirubin levels continue to rise, causing difficulty in distinguishing between physiologic jaundice and HDFN Skeletal growth and muscle development ↑ serum alkaline phosphatase and creatinine levels After age 50, men experience a decrease in secretion rate and concentration of testosterone and women have an increase in pituitary gonadotropins, especially follicle-stimulating hormone (FSH) Gender After puberty, men generally have higher alkaline phosphatase, aminotransferase, creatine kinase, and aldolase levels than women due to larger muscle mass of men Women have lower levels of magnesium, calcium, albumin, Hbg, serum iron, and ferritin. Hemolysis Sheer forces on red blood cells Using too small a needle Pulling the plunger back too fast Expelling blood vigorously into a tube Shaking or mixing tubes vigorously Slight increase in Phosphate, Total Protein, Albumin, Magnesium, Calcium, Alkaline Phosphatase (ALP) Noticeable increase ALT, CK, Iron, Coagulation tests Significant increase: Potassium (K+), Lactate Dehydrogenase (LD), AST Effects of hemolysis on specific analytes Decreases Slight: Haptoglobin, bilirubin Significant decrease in Troponin T The severity of hemolysis must be considered when interpreting values for these analytes

Quality Control

Documentation is EVERYTHING! Ensure the QC sheet is correct and up to date Do NOT report results without ensuring that controls are within expected range Do NOT use expired reagents 3 POC training in a year

Lab Submission Guide

Example FUNGAL SEROLOGIES (IMMUNODIFFUSION (BAMC) 1. Patient Preparation: Aseptic technique. 2. Collection Container: Red top tube. 3. Specimen and Volume Required: 2.5 mL serum. 4. Specimen Processing Instructions: Collect blood in a red top tube, separate serum prior to submission. 5. Cause for Rejection: Less than 2 mL, leaking specimens, or specimens over 24 hours old. 6. Expected TAT: 10 days. 7. Test Performed in Mycology Section, 916-3353. 8. Tests in Panel: HISTO H BAND ID; HISTO M BAND ID; COCCIDIO IDTP ID; COCCIDIO IDCF ID; BLASTOMYCES ID

Indicators of test Reliability/Validity

Four indicators most commonly used to determine the reliability/validity of a clinical laboratory test. Precision and accuracy - How well the test performs day to day Sensitivity and specificity - How well the test is able to distinguish disease from absence of disease. Precision - A measure of repeatability A test method is said to be precise when repeated analyses on the same sample give similar results. When a test method is precise, the amount of random variation is small. The test method can be trusted because results are reliably reproduced time after time Accuracy - A measure of trueness A test method is said to be accurate when the test value approaches the absolute "true" value of the substance (analyte) being measured. Results from every test performed are compared to known "control specimens" that have undergone multiple evaluations and compared to the "gold" standard for that assay, thus analyzed to the best testing standards available. True and false positives - True positive - a person we told is pregnant that really was. True negative - a person we told is not pregnant, and really wasn't. False negative - a person we told is not pregnant, though they really were. Ooops. False positive - a person we told is pregnant, though they weren't. Oh snap! Sensitivity - The ability of a test to correctly identify individuals who have a given disease or condition. A certain test is 90% sensitive If 100 people have a certain disease, the test will correctly identify 90 of those 100 cases (90%). The other 10 people (10%) tested will show a "normal" result which is termed false-negative. The more sensitive a test, the fewer false-negative results will be produced. A negative test result allows you to "SnOut" (sensitivity = rule out) Screening test Specificity - The ability of a test to correctly exclude individuals who do not have a given disease or condition. A certain test is 90% specific If 100 people are healthy, the test will identify as "normal" 90 of those 100 cases (90%). The other 10 people (10%) tested will show a "positive" result which is termed false-positive. The more specific a test, the fewer false-positive results will be produced. A positive test result rules in the target disease ("SpIn"). Confirmatory test A high sensitive test would definitely catch anyone that is positive. So if they're negative, you can be sure that they're negative. A high specificity test would definitely catch anyone that is negative. So if they're positive, you can be sure that they're positive.

Cerebrospinal Fluid

Functions Transport of hypothalamus releasing factors (nutrients) are transported to the cells Provides a mechanical barrier to cushion the brain and spinal cord against trauma. (buoyancy) Provides an excretory waste function (garbage disposal) because the brain has no lymphatic system Protective effect against sudden changes in acute venous (respiratory and postural) and arterial blood pressure or impact pressure Maintains central nervous system ionic homeostasis (blood brain barrier) Cerebrospinal fluid (CSF) is a clear, colorless, sterile fluid that circulates in the ventricles of the brain, the subarachnoid spaces, and the spinal cord Adults produce ~ 500 mL of CSF/day Normal volume 90 to 150 mL of CSF "Turnover" every 5-7 hours CSF is produced mainly by the choroid plexus Absorption of the CSF into the blood stream occurs at the arachnoid villi - "one-way valve" ↑ CSF pressure and ↓ venous pressure, CSF flows into the blood stream ↓ CSF pressure and↑ venous pressure, blood does NOT pass into the ventricular system (BBB)

Therapeutic Drug Monitoring

Goal To increase the likelihood of a therapeutic effect and avoid or minimize adverse effects Must have an appropriate concentration at site of action that produces benefits

Drugs of Abuse

Goal of Drugs of Abuse Testing (DAT) is to detect past exposure or use of a drug Quantitative levels of a drug or its metabolite in fluids are not required It is only necessary to know if the analyte is above (present) or below (absent) a defined cut-off concentration Specimen of choice is almost always urine Point of Care Tests Rapid ID of agent Not for legal purposes Urine Oral fluid

Legal Issues

HIPAA Proper identification of the patient 2 forms of ID Proper labeling of the specimen Patient consent issues Patient privacy issues Chain of custody Blood Alcohol Testing, Toxicology testing

Pre-Collection Variables - Interferences

Hemoconcentration Increases concentration of serum enzymes, proteins, and protein-bound substances, including cholesterol, calcium, and triglycerides, as plasma water leaves the vein due to back pressure. Pt dehydration may also hemoconcentrate Hemodilution When a standing patient reclines, extravascular water transfers to the vascular system and dilutes nondiffusible plasma constituents (20 minutes = 10% decrease)

High Complexity Test

High complexity tests include those that require clinical laboratory expertise beyond normal automation to perform. If the output of the data requires some expertise, these would also be highly complex. Examples include cytology, immunohistochemistry, peripheral smears, flow cytometry, gel electrophoresis, and most molecular diagnostic tests including RT-PCR, gene chip arrays, multiplexed analyses, dot blots, viral loads, expression arrays, etc

Predictive Values

High specificity = few FPs: Sp = TN/(TN+FP);FPs also drive PPV: PPV = TP/(TP + FP);So, the clinician is more certain that a patient with a positive test has the disease (High PPV rules in the disease) The higher the sensitivity = few FN, Sn = TP/(TP+FN); FNs also drive NPV: NPV = TN/(TN+FN); The clinician can be more confident that a patient with a negative score does not have the diagnosis (because there are few false negatives). (High NPV can rule out a disease) We said that before you apply a test, prevalence gives your best guess about the chances that this patient has the disease. This is known as "Pretest Probability of Disease": (TP+FP) / N in the 2 x 2 table: (prevalence) Positive likelihood ratio - rules IN disease LR+ = sensitivity/1-specificity Negative likelihood ratio - rules OUT disease LR- = 1-sensitivity/specificity

Anatomic Pathology

Histology: the study of the microscopic examination of cells and tissues. Cytology: the study of cells in terms of structure, function and chemistry

Provider-Performed Microscopy (PPM)"moderate complexity"

Must be personally performed by a physician, a midlevel practitioner (physician assistant, nurse practitioner, nurse midwife), or a dentist Must be categorized as moderately complex Primary instrument is the microscope The specimen is labile Controls are not available There is limited specimen handling Examples - Direct wet mount preparations for the presence of bacteria, fungi, parasites, and cellular elements for vaginal, cervical, and skin Potassium hydroxide (KOH) preparations Pinworm examinations Fern test Urine sediment examination Nasal smears for granulocytes

Testing for Acetaminophen Toxicity

Overdose of acetaminophen is associated with severe hepatotoxicity Has a high protein-binding affinity Kidney excretion of the drug is minimal Liver damage indicators (elevated enzymes) are usually not detected until 3-5 days after overdose Acetaminophen is rapidly cleared from the serum which makes spot serum testing unreliable as a stand-alone test A nomogram that utilizes a serum acetaminophen level plus a known time interval since ingestion best predicts whether a toxic amount of the drug was ingested Rumack-Matthew nomogram Therapeutic range is 10-30 µg/mL

Procedural Factors

Patient Compliance Patient must take the drug in the proper amount and on a proper dosing schedule to achieve the desired therapeutic level Specimens drawn at the wrong time Blood sample drawn before the drug's steady state is achieved Steady state is achieved when the amount of the drug given equals the amount of the drug excreted by the body Blood sample(s) for trough or peak levels are drawn at an improper time in the dosing cycle The laboratory's test methodology may lack accuracy, precision, sensitivity or specificity

The Clinical Benefits of TDM

Patient non-compliance (under- or over-dosing) can be recognized The patient may have an unusual drug disposition characteristic (noted when the standard dose does not achieve the expected therapeutic effect) Adjustment of prolonged therapeutic drug regimens during periods of continuous physiologic changes Pregnancy or changes that occur as a consequence of an acute or chronic disease

Therapeutic Drug Monitoring

Performed by measuring the concentration of a drug and/or its major metabolite(s) Usually in serum or plasma Using blood levels requires assumptions Proportional relationship exists between plasma/serum concentration and pharmacologic effect For practical reasons blood concentration is measured and not tissue levels where the drug is active Pharmacokinetics provides a mathematical basis to assess the time course of drugs and their effects in the body. It enables the following processes to be quantified: Absorption - Route of administration and formulation of drug will affect rate and extent of absorption Distribution Metabolism Excretion

Phase Contrast Microscopes

Phase contrast is preferable to bright field microscopy when high magnifications (400x, 1000x) are needed and the specimen is colorless or the details so fine that color does not show up well. Effective for observation of cytoplasmic streaming, motility, and the dynamic states of cell organelles Facilitates the study of unstained structures, which can still be alive. Similar to a bright-field microscope, except has changes to the condenser and objective.

Pinworm Examination

Pinworm infection is very common. Female pinworms lay thousands of eggs in the folds of skin surrounding the anus causing itching (bedtime) Touch the perianal skin with transparent tape to collect possible pinworm eggs around the anus first thing in the morning

Specimen Collection

Potential Adverse effects of using wrong amount or improper additive/anticoagulant Interference with the assay Removal of constituents Effect on enzyme action Alteration of cellular constituents Incorrect anticoagulant/blood ratio Put the labels on right so that the individual processing the specimen or the barcode reader won't make mistakes. Yellow has SST which separates the plasma and the RBC after spinning

Analytical Phases of Testing

Pre-Analytical (pre-examination) Provider - order correct test, collect proper specimen Lab - guidance on specimen collection Analytical (examination) Lab - ensure valid result Post-Analytical (post-examination) Lab - provide correct data Provider - properly interpret data →ACTION

Errors

Pre-analytical Error Examples Specimen obtained from wrong patient Specimen procured at the wrong time Specimen collected in the wrong tube or container Blood specimens collected in the wrong order Incorrect labeling of specimen Improper processing of specimen Analytical Error Examples Failure to notice instrument flags Disregard out-of-control quality control results (example on next slide) Wrong assay performed Test procedure incorrectly performed No test performed but a result produced Post-analytical Error Examples Incorrect verbal reporting of alert/critical values Instrument/LIS interface incompatible Confusion about reference ranges Wrong results reported Transposed results during manual data entry Incorrect interpretation leading to mismanagement of patient

Timing of Collection

Prioritization of tests STAT - ER and critical care units 1 hr TAT, highest priority ASAP Hurry up Routine Regular TAT Specific time frame (usually a series of tests): Monitor changes in a patient's condition Cardiac monitoring Determine the level of a medication Peak and trough drug levels The peak specimen is drawn shortly after the medication is given Trough specimens reflect the lowest level in the blood and are generally drawn 30 minutes before the drug is administered. Measure metabolic rate 2-hour postprandial specimen and/or a glucose tolerance test

Analytical

Random vs Systematic Random: Random errors in experimental measurements are caused by unknown and unpredictable changes in the experiment. These changes may occur in the measuring instruments or in the environmental conditions. Systemic: Systematic errors in experimental observations usually come from the measuring instruments. They may occur because there is something wrong with the instrument or its data handling system, or because the instrument is wrongly used by the experimenter.

Common Wet Mount Microscopic Findings

Red Blood Cells: The presence of red blood cells may indicate bleeding during the collection process. In the images below, the arrows indicate examples of red blood cells.

Complete blood count

Red blood cell data Total red blood cell count (RBC) Hemoglobin (Hgb) Hematocrit (Hct) Mean corpuscular volume (MCV) Red blood cell distribution width (RDW) White blood cell data Total white blood cell (leukocyte) count (WBC) A white blood cell count differential may also be ordered Platelet Count (PLT) Total Red Blood Cell Count Count of the number of circulating red blood cells in 1mm3 of peripheral venous blood Normal Male - 4.7 to 5.5 x 106 cells/mcL Female: 3.5 to 5.5 x 106 cells/mcL Hemoglobin Normal Male 13.9-18.0 gm/dL Female 12.2-14.7 gm/dL Critical value < 5 gm/dL or > 20 gm/dL Decrease in Hgb concentration = anemia Increase in Hgb concentration = polycythemia Review value in conjunction with hematocrit concentration Hematocrit is a measure of the percentage of the total blood volume that is made up by the red blood cells Normal Male 39-55% Female 37-48% Abnormal < 25% Caused by anemia, bleeding, RBC destruction, leukemia, malnutrition, overhydration Critical value < 20% or > 60% Hematocrit % = RBC (cells/liter) x MCV (liter/cell) Erythrocyte Indices (calculated from RBC, Hct, and Hgb) MCV (mean corpuscular volume) indicates the average size of the RBCs. Low MCV (microcytic); Normal MCV (normocytic); High MCV (macrocytic) - Helps to diagnose specific type of anemia MCH (mean corpuscular hemoglobin) is the amount of Hgb present in one cell. MCHC (mean corpuscular hemoglobin concentration) is the proportion of each cell occupied by Hgb. MCH and MCHC are discussed together; erythrocytes are normochromic (normal color), hypochromic (less than normal color), or hyperchromic (more than normal color) RDW (RBC distribution width) RDW is an indication of the variation in the RBC size (referred to anisocytosis) WBC - Normal 4,500 - 11,000/mm3 Abnormal Leukocytosis > 11,000/mm3 Caused by infections, leukemia, pregnancy, neoplasms, pneumonia, inflammation or tissue necrosis Leukopenia < 4,500/mm3 Caused by bone marrow failure, radiation, chemotherapy, HIV infection, viral disorders. Critical values < 2,000 or > 30,000/mm3 Platelets - Normal 150,000 to 400,000/uL Abnormal Thrombocytosis > 1 million/uL bone marrow disorder acute bleeding, exercise, heart attack, infections, splenectomy, surgery, etc Thrombocytopenia < 150,000/uL Caused by autoimmune disease, medications, alcohol, viruses, cancer, anemia, etc Critical value < 20,000/uL

Steady-state concentration

Serum drug levels (peak, trough, or random) are most often determined only after steady state has been achieved Assuming first order kinetics, 5 half-lives are required after initiation of drug therapy to reach nearly complete (97%) steady state Five half-lives are also required for near complete elimination after the termination of drug therapy Multiply by 5 to get 5 half lives

Drug Disposition

The key processes involved in drug disposition include liberation, absorption, distribution, metabolism, and excretion, commonly referred to by the acronym LADME. Factors influencing - Demographic Factors Age category (premature infant, neonate, infant, child, adolescent adult, elderly adult) Weight (obesity, malnourishment) Gender Race Other factors influencing - Diseases Related Cardiac diseases may cause a decrease in blood flow GI tract - decreased absorption Receptor tissues - decreased distribution Kidneys - decreased excretion Liver decreased carrier protein levels (affecting drug distribution) results increase the free fraction of the drug in circulation (increased physiologic effect) Absorption of Drug Food or coadministered drug affecting extent and rate of absorption Immediate- or extended-release formulation Gastric pH and motility Distribution of Drug Coadministered drug affecting binding to plasma proteins or tissue receptors Changes in physiologic composition (e.g., rapid weight loss, pregnancy) Metabolism of Drug Food, herbs, or drugs competing for metabolism Coadministration of drug that induces or inhibits metabolic enzymes Excretion of Drug Coadministration of drug that competes for renal tubular secretory paths Changes in urinary flow rate Coadministration of compounds that enhance tubular reabsorption

Evacuated Tubes

Top - Plain Red Contains: No anticoagulant May have a clot activator (thrombin) Yields - Serum Testing area: Chemistry and Serology Top - Red and Gray (SST) "marble" Contains: No anticoagulant May have a clot activator Gel Yields - Serum Testing area: Chemistry Top - Lavender or Purple Contains: Anticoagulant: EDTA Yields - Plasma Testing area: Hematology and Blood Bank testing - Chemistry tests Ammonia, B-type natriuretic peptide (BNP), G6PD, Lead screen Hematology tests CBC, Hematocrit, Hemoglobin, Manual differential, Sickle cell Microbiology tests Blood parasites Top - Light Blue Contains: Anticoagulant: Sodium citrate Yields - Citrated plasma Testing area: Coagulation Coagulation tests - Prothrombin (PT) and international normalized ratio (INR) Partial thromboplastin (PTT) and activated partial thromboplastin time (APTT) D-dimer Fibrinogen Special coagulation tests (factor specific) Top - Green Contains: Anticoagulant: Heparin Yields - Plasma Testing area: Chemistry - Cardiac testing CK, CK-MB, Troponin, Lactate dehydrogenase, Myoglobin, Homocysteine Renal Panel Albumin, Calcium, Carbon Dioxide, Chloride, Creatinine, Glucose, Phosphorus, Potassium, Sodium, Urea Nitrogen Liver Function Tests AST, ALT, LD, AP, TP, Albumin, Bilirubin, Ammonia Toxicology/Therapeutic drug monitoring Top - Gray Contains: Anticoagulant: Potassium Oxalate Glucose preservative (sodium fluoride) TOX draw Yields - Plasma For glycolytic inhibition Testing area: Chemistry - Glucose Testing Glucose, Glucose tolerance test, lactic acid Blood alcohol Lactate Bicarbonate Top - Dark (Royal) blue Contains: Anticoagulant: none, EDTA, or Heparin Yields - serum or plasma Testing area: Special chemistry Used for Trace Metal studies Examples: lead, chromium, cadmium, zinc, arsenic

Lipid Panel

Total cholesterol. This is a sum of your blood's cholesterol content. High-density lipoprotein (HDL) cholesterol. This is sometimes called the "good" cholesterol because it helps carry away LDL cholesterol, thus keeping arteries open and your blood flowing more freely. Low-density lipoprotein (LDL) cholesterol. This is sometimes called the "bad" cholesterol. Too much of it in your blood causes the buildup of fatty deposits (plaques) in your arteries (atherosclerosis), which reduces blood flow. These plaques sometimes rupture and can lead to a heart attack or stroke. Triglycerides. Triglycerides are a type of fat in the blood. When you eat, your body converts any calories it doesn't need into triglycerides, which are stored in fat cells. High triglyceride levels usually mean you regularly eat more calories than you burn. Healthy adults every 5 years - total cholesterol (need not include full panel) If screening cholesterol test result is high, perform the full lipid profile Risk factors other than High LDL Smoking, Age, low HDL, hypertension, family history, diabetes For children and adolescents, routine lipid testing is recommended by the American Academy of Pediatrics (AAP) once between the ages of 9 and 11 and again between 17 and 21. More frequent screening with a lipid profile is recommended for children and youths who are at an increased risk of developing heart disease as adults. Some of the risk factors are similar to those in adults and include a family history of heart disease or health problems such as diabetes, high blood pressure, or being overweight. High-risk children should be tested between 2 and 8 years old, according to the AAP.

Quality system

a formalized system documents processes, procedures, and responsibilities for achieving quality policies objectives Coordinate and direct and organization's activities to meet customer and regulatory requirements Improve effectiveness and efficiency on a continuous basis