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Biology 347
Exam 2 March 27, 2008Name______KEY______
NOTE: Turn off all electronics. Please cover your exam at all time.
I. True/False: (Circle T (true) or F (false) 2 points each, 12 points total)
T F 1. Class II MHC molecules typically bind to longer peptides than do class I molecules.
T F 2. The T cells uses the same set of V, D, and J gene segments as B cells but uses different C gene segments.
T F 3. T cells expressing CD8 recognize antigen (peptide) presented by MHC class I molecules
T F 4. Clonal anergy occurs in T cells if a costimulatory signal is absent
T F 5. Apoptosis is a critical process in the normal functioning of the immune response system.
T F 6. B cell receptor signaling includes membrane expression of immunoglobulin as well as Ig-alpha and Ig-beta molecules.
II. Fill in the blanks (2 points each, 12 points total)
1. Fas/Fas ligand belong to the ____tumor necrosis factor______family of cytokines
2. EGF/EGFR belong to the ______growth factor______family of cytokines
3. Class 1 MHC molecules are expressed on _____most nucleated______cells, expression
of class II MHC molecules are restricted to __dendritic______,
___macrophages______, and ___B______cells.
III. Antigen-antibody reactions. Briefly explain how to conduct each of the immune assays listed below (4 points each, total of 16 points)
a. Indirect ELISA to detect antibodies to HIV in a serum sample
Trap HIV antigens (usually GAG antigens) in plastic wells of 96 well plate. ( Add milk protein or albumin to wells to adhere to plate and prevent antibody from adhering to plate--see if the better students include this, if not, do not count it against them).Wash plate, Incubate plate with serum from individuals suspected of being exposed to HIV, wash and then add enzyme-linked rabbit or mouse antibodies to human Ig. Wash, add substrate and observe for color reaction. Color reaction is a positive test for having antibodies to HIV.
Use your judgment. If they do a competitive ELISA where they lay down a trapping antibody then react antigen with the serum sample then add to well and examine if antibodies in the serum sample blocked antigen from binding to trapping antibody--thiswould work--give some credit since it shows that they know how the ELISA works.
b. Enhanced ChemoLuminescence (ECL) to detect MHC class I molecules (45 kd) in a cell membrane preparation
Perform western immunoblot with solubilized membrane preparation. ( Next, transblot proteins to matrix such as nitrocellulose---see if students get this, if not, don't count it against them). React transblotted paper with primary antibody to class I proteins. Next, add horse radish peroxidase-linked secondary antibody + luciferin and oxidizers and expose to x-ray film in the dark. Light emitted will expose film, indicting where class I molecules are located. Looking for a protein of 45kd in size.
c. ELISPOT to detect T cells secreting IL-4
Plate cells on plastic that has IL-4 trapping antibody. Remove cells (careful attention where cells are located). Next, add secondary antibody with enzyme. Add insoluble substrate and examine for color reaction where cells were previously plated.
d. Immunefluorescence to determine the ratio of T helper to cytotoxic T cells in a blood sample
Culture blood cells with fluorescent (green or red) tagged CD4 (red) or CD8 (green). Use fluorescent activated cell sorter to sort (count) the number of red and green cells. Can also use fluorescent microscope to manually count cells.
IV. Short Answers/diagram/definitions (total of 50 points. (Be concise and answer in space provided)
1. Define the following terms (3 points each, 15 points total)
a. MHC self restricted
Thymus development of T helper and cytotoxic T cells that are selected on the basis that they recognize self (at low affinity binding). For both cell types to become activated they must recognize self and then altered self or self expressing foreign antigen.
b. Antigen processing
Processing of exogenous proteins by antigen presenting cells. For cytotoxic T cells most nucleated cells process proteins into peptides of approximately 13-18 amino acids and present via class I proteins. For T helper cells (B cells,dendritic cells,macrophages--count tissue dendritic cells correct if students list them) digest proteins into peptides of 8-10 amino acids and present to T helper cells via class II proteins.
c. Haplotype
The set of alleles of linked genes present on one parental chromosome; commonly used in reference to MHC (class I and II MHC genes).
d. Thymus dependent antibody response
Amplified immune response involving affinity maturation, isotype switching, and generaton of memory cells. (some students may draw primary and secondary antibody response showing IgM and IgG levels)
e. Superantigens
Protein from virus, bacteria, parasite that has the ability to bind to APC and T cell with high affinity, thereby activating T cell in the absence of peptide--i.e, no specificity
2. Briefly describe the developmental events that occur at each stage of B cell differentiation/maturation. Indicate which events occur in the bone marrow or in the periphery--germinal centers (3 points each, 15 points total)
a. Pro-B cell (occurs in bone marrow)
Expression of Ig-alpha/Ig-beta receptor on membrane (expressed through out B cell development)
D-J joining of mu (IgM) heavy chain
b. Pre-B cell (occurs in bone marrow)
V, D, J, C (mu) giving rise to mu heavy chain expressed on membrane. Heavy chain binds to surrogate "light chain like" protein and under goes cell proliferation, a single cell generating approximately 250 pre-B cells
c. Immature B cell (occurs in bone marrow)
VJC joining to generate kappa or lambda light chain. Establish B cell tolerance by eliminating (apoptosis) cells where IgM binds to self antigens. A small percentage of cells escape apoptosis by editing light chains which involves activation of RAG1 and 2 and replacing VJ regions of light chains such that the IgM does not interact with self antigens
d. Naive B cell (lymph nodes)
Migrate to lymph nodes--germinal centers, follicles, naive in that they have not been exposed to antigens of pathogens
e. Mature B cell (lymph node--germinal center--follicles)
exposed to antigens of invading pathogens. Ig undergoes affinity maturation via mutation by somatic mutations in V region hypervariable regions, undergo class switching and generation of memory B cells.
3. Diagram and label the various molecules involved in the interaction of a B cell with a T helper cell, resulting in the activation of the T helper cell and proliferation of the B cell (8 points total)
page 239 of text
4. Explain or diagram positive and negative selection of T cells in the thymus (6 points total)
page 250 of text
V. Multiple Choice (2 point each, 10 points total)
1. A thymus independent antigen will not elicit an immune response involving
A. isotype switching
B. affinity maturation
C. generation of memory B cells
D. A and C are correct
E. A, B, and C are correct
2. Explain the biological consequence of treating fibroblasts and epithelial cells with the cytokine transforming growth factor beta (TGF-beta)
A. growth inhibition of fibroblasts as well as epithelial cells
B. growth activation of fibroblasts as well as epithelial cells
C. growth activation of fibroblasts and growth inhibition of epithelial cells
D. growth inhibition of fibroblasts and growth activation of epithelial cells
E. none of the above
3. Treat immature B cells with an antibody that specifically reacts with membrane expressed IgM will cause the immature B cells to:
A. undergo Ig isotype switching
B. differentiate into mature B cells
C. proliferate
D. undergo apoptosis
E. none of the above
4. Lipopolysachride (LPS) is can be referred to as a
A. polyclonal activator of B cells
B. B cell mitogen
C. T cell independent antigen
D. B and C are correct
E. A, B, and C are correct
5. A mechanism for generating antibody affinity maturation is:
A. light chain editing
B. heavy chain editing
C. substitution of V, D, or J regions
D. somatic mutations in hypervariable regions
E. isotype switching
6. Cytokines can exhibit
A. pleiotrophic effects
B. redundancy
C. synergistic effects
D. antagonistic effects
E. A, B, C, and D are correct
7. Transcription factor that is a key player in immune cell proliferation
A. SP1
B. SP3
C. NFkB
D. FADD
E. BID
8. The interleukin-2 (IL-2) receptor can function as:
A. monimer
B. dimer
C. trimer
D. A and B are correct
E. A, B, and C are correct