You are studying the immune response to Shigella in mice. You infect the mice wi
ID: 71906 • Letter: Y
Question
You are studying the immune response to Shigella in mice. You infect the mice with bacteria for one week. Following infection, you sacrifice the mice and isolate white blood cells from the spleen. If the mice are mounting an immune response then CD4+ T cells should be actively dividing and entering cell cycle. You take the total splenocytes (spleen cells) and culture them in the presence of BrdU and bacteria. Then you harvest the cells and stain them for BrdU and CD4 and analyze by flow cytometry. Interpret the results below. Are there CD4+ T cells present? Have the CD4+ T cells recently gone through S phase?Explanation / Answer
Splenocytes consist of different cell types including CD4 and CD8 cells. In the figure the fluorochrome used to stain CD4 cells is APC.
The figure represents 4 quadrants
Lower left: Indicates negative for BrdU staining and CD4
Lower Right: Positive for CD4 and negative for BrdU
Upper left: Positive for BrdU and negative for CD4
Upper right: Positive for CD4 and negative for BrdU
Coming to the first question, the answer is CD 4 T cells are present. You can see the cluster of cells in the lower right quadrant in both the graphs. It gives an indication of CD4+ T cells in the splenocytes.
Coming to the second question, BrdU is incorporated into the actively growing cells during the S-phase of the cell cycle substituting for thymidine. Antibody against the BrdU is used, so any signal from BrdU gives an indication that cells have recently gone through S-phase.
From the first graph, it is clear that you cannot get any positive BrdU signal because there is BrdU.
From the second graph, there are BrdU positive cells (upper right and left quadrant) which are both CD4 T cells and other cells in the splenocytes. So the answer is CD4 T cells have recently gone through S-phase (Upper right quadrant).
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