Deck 20: Experimental Methods and Systems

Question 1

Polyclonal antibodies differ from monoclonal antibodies in all ways EXCEPT which of the following?

A)They generally have fewer cross-reactivities.
B)They are generally better for immunoprecipitation.
C)They generally have lower affinity.
D)They can vary from preparation to preparation.
E)All of the answers are ways in which they differ.

Answer 1

A

Question 2

Monoclonal antibodies make effective research tools because

A)as products of a single B cell, they all have the same specificity.
B)their specificity is stable over time.
C)their cross-reactivity can be well characterized.
D)they are produced by transformed cells and, therefore, can be produced in large quantities.
E)All of the answers are correct.

Answer 2

E

Question 3

Which of the following is NOT required to generate a B-cell hybridoma?

A)Myeloma cell line
B)Primary plasma cell
C)Chemical fusogens
D)HGPRT selection
E)None of the answers are required to generate a B-cell hybdridoma.

Answer 3

E

Question 4

Immunoprecipitation does NOT

A)allow characterization of molecules bound to cells.
B)take place in gel matrices.
C)work well in solution.
D)require the use of a monoclonal antibody.
E)All of the answers are characteristics of immunoprecipitation.

Question 5

Immunoprecipitation can be used

A)with Western blotting to assess its efficiency.
B)with Western blotting to determine protein abundance.
C)with secondary antibodies to ascertain protein-protein interactions.
D)to purify proteins.
E)All of the answers are correct.

Question 6

All of the following concerning agglutination reactions are true EXCEPT

A)they are used only with red blood cells.
B)they only detect molecules on the surface of cells.
C)they require a high degree of expertise to interpret.
D)they should be done with titrations of antibodies.
E)All of the answers are true.

Question 7

Hemagglutination inhibition reactions are used to detect

A)responses to bacteria.
B)responses to viruses.
C)proteins on the surface of red blood cells.
D)All of the answers are correct.
E)None of the answers are correct.

Question 8

ELISPOT assays

A)are modifications of Western blotting.
B)are used to detect individual cells.
C)involve the use of "capture" antibodies.
D)both involve the use of "capture" antibodies and are modifications of Western blotting.
E)both involve the use of "capture" antibodies and are used to detect individual cells.

Question 9

Design of an effective ELISA can be challenging because

A)different detection systems have sensitivity that ranges over five orders of magnitude.
B)the number of replicates required for any ELISA is quite high.
C)the anticipated concentration of targets must be estimated to determine the sensitivity required of the assay.
D)signal amplification by the assay conditions can impair interpretations.
E)All of the answers are correct.

Question 10

Which of the following does NOT describe equilibrium dialysis?

A)It is used to quantitate binding between antibody and antigen.
B)It depends on the ligand being able to cross a semipermeable membrane.
C)It measures the amount of antibody-bound ligand compared to unbound ligand.
D)It requires an enzyme amplification step.
E)All of the answers describe equilibrium dialysis.

Question 11

Surface plasmon resonance

A)is a relatively recently developed technique.
B)depends on the molecules close to a reflective surface.
C)requires that both binding and dissociation be evaluated.
D)All of the answers are correct.
E)None of the answers are correct.

Question 12

Immunohistochemistry and immunocytochemistry do NOT share which of the following characteristics?

A)Use of antibodies to recognize specific targets.
B)Use of enzymes to amplify the signal.
C)Use of irrelevant proteins, such as milk, to reduce nonspecific binding.
D)Use of detergent to permeabilize the membrane.
E)They do share all of these characteristics.

Question 13

Immunohistochemistry and immunocytochemistry differ from each other in that

A)immunocytochemistry analyzes tissue sections.
B)immunohistochemistry analyzes individual cells.
C)immunohistochemistry uses electron-gold microscopy.
D)Immunocytochemistry analyzes individual cells.
E)None of the answers are correct.

Question 14

Immunofluorescence-based imaging

A)allows fine-scale visualization of subcellular localization.
B)has been enhanced by the availability of multiple dyes.
C)has been simplified by genetic engineering to tag GFP onto target proteins.
D)can take advantage of molecules that do not need antibodies to bind targets.
E)All of the answers are correct.

Question 15

Confocal microscopy improves on immunofluorescence in that it

A)increases the resolution of the image.
B)allows construction of three-dimensional images.
C)allows detection of multiple molecules.
D)is the only technique that allows use of living cells.
E)None of the answers are correct.

Question 16

Multiphoton microscopy is a variation on what type of technique?

A)Confocal microscopy
B)FACS
C)MACS
D)Electron microscopy
E)Immunohistochemistry

Question 17

In flow cytometry, in addition to fluorescent markers, forward (FSC) and side (SSC) scatter are measured, and

A)side scatter is a reflection of the size of the cell.
B)forward scatter is a measure of the amount of DNA in the cell.
C)side scatter is a measure of the amount of DNA in the cell.
D)forward scatter is a reflection of the internal complexity of the cell.
E)None of the answers are correct.

Question 18

Intracellular staining is a major advance in cell sorting that

A)requires permeabilization of the cell membrane.
B)allows detection of molecules inside the cell instead of simply at the cell surface.
C)allows subcellular localization of molecules.
D)both requires permeabilization of the cell membrane and allows detection of molecules inside the cell instead of simply at the cell surface.
E)both requires permeabilization of the cell membrane and allows subcellular localization of molecules.

Question 19

The major distinction between FACS and MACS is that

A)MACS is not as precise as FACS.
B)MACS does not use a fluorescent marker.
C)MACS can be used with living cells.
D)MACS can be used for intracellular staining.
E)All of the answers are correct.

Question 20

You might want to use MACS instead of FACS if you

A)have a very small number of cells to sort.
B)are working with cells that are especially fragile.
C)are working with a very large number of cells.
D)need a very high degree of accuracy.
E)are using multiple antibodies.

Question 21

Which of the following is TRUE regarding propidium iodide and cell cycle analysis?

A)Apoptotic cells will have more DNA than normal G1 content.
B)S-phase cells will have the same DNA content as G1 cells.
C)G2 cells will have more DNA than normal G1 cells.
D)M cells will have the same DNA content as normal G1 cells.
E)All of the answers are true.

Question 22

Carboxyfluorescein succinimidyl ester (CFSE) can be used to track cell division because

A)after pulsing the cells with it, dividing cells will incorporate more into their DNA.
B)after pulsing cells with it, dividing cells will incorporate more into their proteins.
C)its intensity is cut in half, approximately, when DNA replicates.
D)its intensity is cut in half, approximately, when the cell divides.
E)None of the answers are correct.

Question 23

In chromium release assays, or more recently in CFSE release assays, cells that die release the marker if they

A)die by apoptosis only.
B)die by necrosis only.
C)die by either apoptosis or necrosis.
D)divide before dying.
E)replicate DNA before dying.

Question 24

Annexin-V staining of cells identifies

A)a failure of membrane integrity.
B)broken ends of chromosomes produced during DNA fragmentation.
C)necrotic but not apoptotic cells.
D)both apoptotic and necrotic cells.
E)a breakdown of membrane asymmetry.

Question 25

The following can be used to analyze chromatin structure in intact cells.

A)flow cytometry
B)ChIP
C)3C
D)3C and ChIP only
E)3C and flow cytometry only

Question 26

CRISPR-Cas9 technology can be used to

A)remove targeted sequences of DNA.
B)induce recombination and introduction of mutations.
C)repair genetic diseases.
D)tag specific DNA for imaging.
E)All of the answers are correct.

Question 27

The use of inbred lines of mice

A)reduces genetic heterogeneity.
B)reduces experimental variables.
C)increases similarity among individuals.
D)leads to syngeneic animals.
E)All of the answers are correct.

Question 28

Congenic strains are NOT

A)identical genetically at almost all loci.
B)distinct at only one locus.
C)useful for adoptive transfer experiments.
D)equivalent to identical twins.
E)All of the answers are congenics.

Question 29

The Cre/lox system is NOT able to

A)delete a gene in a specific tissue type.
B)delete a gene when a drug is introduced.
C)express a gene only in specific tissues.
D)enable study of embryonic lethal gene alterations.
E)All of the answers are correct.

Question 30

How do monoclonal and polyclonal antibodies differ from one another? Explain, and provide two advantages in using each type of antibody in assays.

Question 31

Compare and contrast indirect and sandwich ELISA assays.Which requires multiple types of antibodies against the target antigen? Explain.

Question 32

Explain how an ELISPOT works.What does it most commonly measure?

Question 33

What are two methods used to measure the affinity of antibodies to antigen? Which method is newer? Explain.

Question 34

What are two useful techniques for visualizing the intracellular location of a protein in groups of cells in intact tissues? What links them? Explain.

Question 35

You are studying the localization of stored granzyme and an internal membrane marker in activated T cells.How could you image groups of these two molecules together when they are located all over the 3-D space inside of cells? Explain.

Question 36

What simpler technique from immunology is the basis for modern flow cytometry, and how does it improve on the earlier assays? Explain.

Question 37

List and evaluate three ways to measure cell death experimentally.

Question 38

Using what you have learned in the text, explain the excitement surrounding the technological innovation(s) behind CRISPR-Cas9.In your answer, describe how CRISPR-Cas9 works.

Question 39

Imagine you want to study Promoter X, which controls many of the genes in B cells that you believe are important for controlling viral infections.You want to generate a transgenic mouse line with a mutation in this promoter that makes it expressed at low levels.Unfortunately, after all of your work, you learn that your mutation is "embryonic lethal." Describe the strategies/techniques that could be used to circumvent this problem.