Question

24) 100000 10000 CD4 PE اما مقيماملی 0 100 100000 1000 10000 CD8 FITC 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? C) 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 T cells that are found within quadrant 3.

Answer 1

Answer A: Quadrant-3

Comments: The developing T cells are double-negative (DN) and they do not express CD4 or CD8 markers on its cell surface. So this type of cells in flow cytometry data analysis lines in Quadrant-3.

Answer B: Quadrant-2

Comments: The double-positive T cells (DP) are expressing both CD4 and CD8 on its cell surface. In flow cytometry data analysis the double-positive cells line in Quadrant-2.

Answer C: Quadrant-1

Comments: The MHC-II binding T cells are CD4⁺. Further the CD4⁺ T cells in flow cytometry data analysis line in Quadrant-1.

Answer D: CD45 and CD3

Comments: The T cells are developed in bone marrow through hematopoietic stem cells (HSCs). Initially the HSCs cells are expressing CD34; during the development the sequential expression of CD45, CD3, CD4 and CD8 takes place on the cell surface of T cells. So in the initial developing phase of T cells, these cells are negative for CD45 and CD3 and these types of cells in flow cytometry data analysis lie in Quadrant-3.

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