Cell Bio Exam 3

A standard curve is prepared by making serial dilutions from a known concentration of the antigen. This yields a gradient from high to low (or low to high depending how the standards are plated). With a colorimetric detection method, this would give a decreasing intensity of the color (or increasing if the concentration goes low to high). Which columns are most likely the standards?

1 and 2

Based on this graph if you had a sample with an OD of 1.0, the concentration of GRP78 would be approximately...

10 ng/ml

For colorimetric detection, what equipment is used? What does it measure?

A color is produced by the product to guess if the antibody/antigen is present. No color means negative and yellow means antigens/antibodies are present. This is done by a serial dilution or kit. A standard curve is done with known concentrations of antigens/antibodies to quantitate how much protein is present. A plate reader is needed to read absorbance and optical density.

What is the signal molecule involved in contact-dependent signaling?

A membrane ligand (protein, oligosaccharide, lipid) and a membrane protein of two adjacent cells interact

How is a standard curve created?

A standard curve is created by performing a serial dilution, which provides the standards (concentrations) with which to plot a standard curve. The standards are ran through a spectrometer where an optical density is obtained for each. The concentration of your protein in each well (x-axis) is plotted against the optical density (y-axis). Then a curve is drawn through the points, showing what optical density can be expected at different concentrations, and vice versa.

What is the function of ARP? What does ARP stand for?

ARP stands for Actin Related Protein The function of ARP's is to control the behavior of actin filaments in vertebrate cells. They bind to theses filaments and hold them together. They also allow for branching to occur so were not just getting filaments in a straight linear direction.

What is the signal molecule involed in neuronal signaling?

Acetylcholine and y-Aminobutyric Acid (GABA)Both are used to signal neurons down the chain

Name two types of enyzme sytems used for ELISAs.

Alkaline phosphatase (AP) and Horseradish peroxidase (HRP) are two available enzyme systems for ELISA assays.

Would this antibody, anti-Sos antibody conjugated HRP, be used as a primary or secondary antibody?

Anti-Sos antibody conjugated HRP would be used as a secondary antibody because it is conjugated which would allow us to use various techniques to detect the antigens that it is connected to.

What is the function of the GTP cap?

At the 5' end, a cap is added consisting of a modified GTP (guanosine triphosphate). This occurs at the beginning of transcription. The 5' cap is used as a recognition signal for ribosomes to bind to the mRNA.

You purchase an IL-6 ELISA kit. The kit arrives and the plate is not pre-coated. You have two antibodies in the kit. One is the primary/coating/capture antibody; the other is the secondary/detector/detecting antibody. You also found one from a previous ELISA kit you had purchased. Select the antibody that is the primary antibody A) Biotin-Conjugate anti-mouse IL-6 monoclonal antibody B) Anti-mouse IL-6 polyclonal antibody C) HRP-Conjugate anti-mouse IL-6 monoclonal antibody D) None of the above

B) Anti-mouse IL-6 polyclonal antibody

Which of the wells listed below would be considered strong positives? Strong positive will be equal to or great than C1 and C2 standards.

B11 and B12

An actin-binding protein called cofilin binds preferentially to ADP-containing actin filaments rather than ATP-containing actin filaments. Based on this preference, which is true? A) Cofilin binds to the plus ends of treadmilling actin filaments. B) Cofilin competes with profilin for binding to actin. C) Cofilin binds to older actin filaments. D) Cofilin binds to the plus ends of actin filaments.

C) Cofilin binds to older actin filaments.

Which does not contain both actin and myosin? A) a contractile bundle in a nonmuscle cell B) a muscle cell sarcomere C) the lamellipodium at the leading edge of a crawling cell D) the contractile ring that carries out cytokinesis

C) the lamellipodium at the leading edge of a crawling cell

T or F: For cell signaling, receptors must be on the plasma membrane. Signaling molecules are not able to enter the cell on their own

Cell-surface receptors are membrane-anchored proteins that bind to ligands on the outside surface of the cell. In this type of signaling, the ligand does not need to cross the plasma membrane. So, many different kinds of molecules (including large, hydrophilic or "water-loving" ones) may act as ligands

Which type of ELISA only uses one antibody?

Direct ELISA Substrate is then added, producing a signal that is proportional to the amount of analyte in the sample. Sinceonly one antibodyis used in a directELISA, they are less specific than a sandwichELISA. When toUse: Assessingantibodyaffinity and specificity.

What type of approach is an ELISA? Biochemical? Genetic? Microscopy?

Elisa is a biochemical approach.

What does ELISA stand for?

Enzyme Linked ImmunoSorbent Assay ELISA is a test that detects and measures antibodies in the bloods, and is a reliable way detect harmful proteins that the body produces

T or F: Beta-galactosidase or beta-gal is a commonly used enzyme system for ELISA assays

False

T or F: G-protein coupled receptors (GPCR) are single pass transmembrane proteins

False

T or F: Kinesin moves vesicles in retrograde transport

False Nearly all kinesins move cargo toward the (+) end of microtubules (anterograde transport), whereas dyneins transport cargo toward the (−) end (retrograde transport). ... Each type of membrane vesicle is transported by its own kinesin motor protein.

T or F: The same signal molecule can only induce the same response in different target cells

False The same signal molecule can induce different responses in different target cells. An image that shows this is also found on the same slide

Why are fluorogenic and chemiluminogenic not as common detection methods?

Fluorogenic gets fluorescence but requires an extra step and more equipment to detect this. Chemiluminogenic is when light gives off radiation and this also requires an extra step and equipment. It also has a short read out time meaning you have to work quick. Colorimetric detection is the most common method because all you are looking for is a color change and does not require equipment.

Dynamic instability in microtubules stems from the intrinsic capacity of tubulin molecules to hydrolyze what?

GTP

When a signal needs to be sent to most cells throughout a multicellular organism, the signal most suited for this is a ___________.

Hormone

How do GTP and ATP play a role in the cytoskeleton?

How do GTP and ATP play a role in the cytoskeleton? GTP can be found in tubulin monomers, and can hydrolyze in the process growing longer microtubules. Remember that the plus end grows faster than the minus end, and treadmilling is a process that can also occur during the growth process. ATP can be found in the actin monomers, and phosphorolyze in the process of growing longer actin filaments. Again there is a plus end that grows faster than the minus end, and treadmilling can occur during the growth process as well. Microtubules are responsible for cell division and cell transport within the cytoskeleton, and actin is responsible for different cell surface projections within the cytoskeleton.

If you want to detect antigen/protein what type of ELISA is best?

If you want to detect a antigen or protein you would use a sandwich ELISA because the sandwich ELISA detects presences and concentration of antigen/protein.

If you want to detect antibody what type of ELISA is best?

If you want to detect antibody, you will used the indirect ELISA because it detects presence and concentration of antibody in a sample

How are ELISAs used in health care?

In the healthcare environment ELISAs can be used in testing for antibodies. Specifically, an indirect ELISA will be used to detect the presence of antibodies for a virus, bacteria, or a different pathogen from a sample. The results would be indicated via a change in color of the ELISA's solution. This same process with the ELISA is also used in pregnancy tests.

Match the type of ELISA with what it is used to detect detects presences and concentration of antigen/protein detects presences and concentration of antibody

Indirect ELISA: detects presences and concentration of antibody sandwich ELISA: detects presences and concentration of antigen/protein

What does "9 +2" refer to?

It's a peripheral ring of 9 microtubular structures and 2 central microtubules attached to the 9 outer microtubules carry two rows of dynein molecules which move like a ratchet giving it its force to beat the cilia

Name a drug that affects actin filaments

Latrunculin effects actin filaments by binding to actin monomers, preventing their polymerization.

What is the function of the MTOC? What is another name is called?

MTOC stands for Microtubule Organizing Center. It has two main functions: The organization of eukaryotic flagella and cilia. The organization of meiotic and meiotic spindle apparatus that separate chromosomes during cell division.

What is the subunit that makes up microtubules?

Microtubules are the largest type of filament, with a diameter of about 25 nanometers (nm), and they are composed of a protein called tubulin. Actin filaments are the smallest type, with a diameter of only about 6 nm, and they are made of a protein called actin.

What is the function of microtubules?

Microtubules function as transportation highways to move organelles around the cell. They are like conveyer belts!

Which type of filaments have a plus and minus end? What is the difference between plus and minus end?

Microtubules have a plus and minus end. The difference between the two is that there is faster growth at the plus end of the microtubule as compared to the minus end. Another difference is the minus end is called the pointed end while the plus end is called the barbed end.

What molecule is need to allow for motor protein to "walk"?

Motor proteins fulfill the role of transporting large cargo about the cell to their required destinations. Kinesins are motor proteins that transport such cargo by walking unidirectionally along microtubule tracks hydrolysing one molecule of adenosine triphosphate (ATP) at each step.

What is the motor protein associated with actin?

Myosin are motor proteins that are usually found interacting with actin

What is the signal molecule involved in paracrine signaling?

Paracrine signaling molecules are called local mediators because they act on nearby cells. Examples of these molecules are listed below. Nitric Oxide (NO) gas in nerve cells and endothelial cells. Histamine in mast cells. Platelet-derived growth factor in various cells including blood platelets.

What is a sandwich ELISA?

Sandwhich ELISA measures Ag presence and levels. It detects presence and concentration of antigen/protein levels.

What is a basal body? Where is it found?

Stable microtubules, arranged in a bundle grow from a cytoplasmic basal body, which serves as an organizing center. Microtubles found at the base of cilia and flagella are called basal bodies.

Name two cell structure made up for actin that allows for movement

Stress Fibers: contract and help the cell to shift or have more flexibility. Filopodia: Pseudopodia (Amoba) and Filopedia both interchangeable and help with movement

Name a drug that affects microtubules

Taxol is a drug that affects microtubules, by binding to and stabilizing them. Colchicine and Vinblastine

Pick a signaling pathway. What is its signal molecule, receptor, and cell response?

The G-protein coupled receptor is a signaling pathway, also known as 7 transmembrane proteins and uses phosphorylate C and Adenylate cyclase as a signal molecule. The receptor is a G-protein which consist of 3 subunits alpha, beta, gamma. Cell Response: 1) Ligand binds to GPCR 2) GPCR undergoes conformational change 3) Alpha subunit exchanges GPP for GTP 4) Alpha subunit dissociates & regulates target proteins 5) Target protein relay signal via 2nd messenger 6) GTP hydrolyzed to GDP

What are the three filaments that make up the cytoskeleton? List them from smallest to largest

The actin, intermediate filaments, and microtubules are the three filaments that make up the cytoskeleton. From smallest to largest: Actin (5-9nm diameter), intermediate filaments (10nm diameter), microtubules (25nm diameter).

Would this antibody, anti-Sos antibody, be used as a primary or secondary antibody?

The anit-Sos antibody would be used as a primary antibody. It is used for the direct detection of an antigen, It may produce some false positives.

What is the function of plectin?

The function of plectin is to enable the interaction between filaments by cross linking them to different networks of structural proteins.

What is the function of intermediate filaments?

The role of intermediate filaments is mechanical strength in the cytoskeleton

What is the signal molecule involved in endocrine signaling?

The signal molecule involved in endocrine signaling is hormones

What is the subunit that makes up actin filaments?

The subunit that makes up actin filaments is an actin monomer.

What type of subunits make up intermediate filaments?

The type of subunits that make up intermediate filaments are staggered long subunits.

What does treadmilling refer to?

Treadmilling is a phenomenon observed in many cellular cytoskeletal filaments, especially in actin filaments and microtubules. It occurs when one end of a filament grows in length while the other end shrinks resulting in a section of filament seemingly "moving" across a stratum or the cytosol.

T or F: ELISAs are used to detect antibodies in patients' blood to determine if they have be exposed and developed an immune response to a pathogen, such as SAR-CoV-2 (COVID-19), influenza, Borrelia burgdorferi (Lyme disease), etc. ELISAs can also be used to detect antibodies for allergies, such as food allergies

True

T or F: Fluorogenic and chemiluminogenic require specialized equipment so there are not a common detection method

True

T or F: Some signal molecules can bind directly to intracellular proteins that bind DNA and regulate gene transcription

True

T or F: Washing the plate in between each step is important to remove unbound and excess material i.e. proteins in the lysate that are not the antigen/protein of interest, excess antibody that did not bind the antigen, etc.

True

T or F: The response by a cell depends on the multiple signals it receives

True the cells response is dependent on the signals because they tell the cell whether to survive, die or grow and divide.

What is an indirect ELISA?

When describing an indirect ELISA it is important to note that contrast to an uncommon direct ELISA, an indirect ELISA utilizes a Secondary Antibody Conjugate. This utilization is an attempt to detect the presence and concentration of antibodies in a sample. When conducting this method one will receive a plate that has an Antigen-coated well, then after this one will need to perform a was and add specific antibody to which will be measured. Another wash follows this, and one will need to add an enzyme-conjugated secondary antibody. Lastly, after another wash, one will add a substrate and then measure its color. Ultimately, this method is used to appropriately measure antibodies.

Match the letter with the level of cell signaling

__A__ extracellular signaling molecule __D__ cell responses __B__ effector proteins __E__ receptor protein __C__ intracellular signaling molecules

Match the type of ELISA with its image

__C__ sandwich ELISA __A__ direct ELISA __B__ indirect ELISA

What is the function of actin filaments?

actin filaments are used for structural purposes, cell mobility and help function other proteins like myosin for muscle contraction.

Which is the toughest and most durable of the different types of cytoskeletal filaments? HINT: They stop the cell from being ripped apart.

intermediate filaments

Which of the following cytoskeletal structures are the most common for providing tracks for guiding intracellular transport?

microtubules

What does the cellular motility of sperm depend on?

microtubules and dynein

What are small intracellular signaling molecules often called?

second messengers

When cells respond to an extracellular signal, they most often convert the information carried by this molecule from one form to another. What is this process called?

signal transduction