||anti-CD3, anti-CD28 activated CD4 positive T cells
||Annexin V staining was performed by washing CD4 positive T cells with BioLegend staining buffer, resuspending them in Annexin V Binding buffer and then staining with APC Annexin V (1:20). After 15 min of incubation time at room temperature in the dark, the cells were washed and resuspended in Annexin V Binding Buffer for flow cytometric measurement.
||The APC Annexin V antibody works very well on activated T helper cells. CD4+ sorted T cells were activated with aCD3/aCD28 for 48 hours and then stained with CD4 FITC, 7-AAD and Annexin V APC. The graph shows the gating strategy: gating on lymphocytes (P1) and single cells (P2), then stained for CD4 positive cells (P3). Finally, viability of Th cells was assessed using 7-AAD and Annexin V staining (P4), showing that 35% of cells were pre-apoptotic and 8% were double positive for 7-AAD and Annexin V.