Question

Immunolgy 1 question Please answer

100000 10000 CD4 PE 1000 100 0 100 100000 1000 10000 CD8 FITC (5 pts) The above flow cytometry profile was obtained from staining thymocytes with anti-CD4 and anti-CD8 antibodies. Answer the following questions based on your combined understanding of how to read flow cytometry profiles together with an understanding of the events occurring in T cell development and selection. A) In which quadrant are the developing T cells undergoing VDJ recombination? B) In which quadrant is both positive selection and CD4 versus CD8 lineage commitment occurring? ) In which quadrant do you find developing T cells that express TCRs that bind to antigens presented in MHCII? D) What 2 additional antibodies specific to surface proteins would allow identification of the 4 populations of developing I cells that are found within quadrant 3.

Answer 1

A. VDJ rearrangements occur in double negative T cells that lack CD4 and CD8 antigens. These DN cells are of four types- DN1, DN2, DN3 and DN4 based on expression of CD44 and CD25. These cells undergo recombination of Tcrd, Tcrg and Tcrb genes These DN cells are present in quadrant 3. These cells in quadrant 3 will lack CD4 and CD8 expression.

Right choice: Quadrant 3.

B. The cells that undergo positive selection will express both CD4 and CD8 antigens. The cells that do not undergo this beta selection or positive selection will be lost by apoptosis. The quadrant 2 has cells that express both CD4 and CD8 antigens. These cells will then interact with MHC class I or Class II molecules in the cortex. However, only those cells that show strong interaction will only survive and will then migrate to medulla to undergo negative selection. This negative selection will then select for cells that react with self-antigens. Later on there will be separation into CD4+ and CD8+ cells. Thus, quadrant 2 will have cells undergoing positive selection and CD4 vs CD8 lineage commitment.

Right choice: Quadrant 2.

C. T Cells that bind to MHCII (Class II MHC) should express CD4 antigens. T Cells that express CD8 molecules will bind to antigens that are presented with Class I MHC molecules. Quadrant 1 has T cells that express CD4 antigens. Quadrant 4 has T cells that express CD8.

Right choice: Quadrant 1.

D. The four populations of double negative T cells express either CD44 and/or CD25 antigens. DN1 are CD44+CD25-, DN2 are CD44+ CD25+, DN3 are CD44- CD25+ while DN4 are CD44-CD25-. CD3 antibody will detect all T cells.

Right choice: CD25 and CD44 antibodies.