Fixable Viability Stain 575V

Flow cytometric analysis of human Jurkat cells stained with BD Horizon™ Fixable Viability Stain 575V. Cells from the Jurkat (Acute T cell leukemia, ATCC TIB-152) cell line were treated with 0.025% DMSO (Top Left Panel) or 5 μM camptothecin (Top Right Panel) for 16 hours and then stained with BD Horizon™ Fixable Viability Stain 575V (FVS575V; Cat. No. 565694) in serum-free buffer. The cells were then washed and either left unfixed (solid line histograms), or fixed in BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized in BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885) (dashed line histograms). The FVS575V-stained, fixed and permeabilized cells were further stained with Alexa Fluor® 647 Rabbit Anti-Active Caspase-3 antibody (Cat. No. 560626) to identify apoptotic cells (Bottom Panels). Based on co-staining with Anti-Active Caspase-3 and FVS575V, live Jurkat cells are double negative, apoptotic Jurkat cells are Active Caspase-3-positive and FVS575-dim to mid-positive, and dead Jurkat cells are Active Caspase-3-positive and FVS575-positive. Histograms and contour plots were derived from gated events with the forward and side light-scattering characteristics of intact Jurkat cells. Flow cytometric analysis was performed using a BD LSR II Flow Cytometry System. Please note that FVS-575V is also compatible with BD Phosflow™ Perm Buffer III (Cat. No.558050) or BD Pharmingen™ Transcription Factor Buffer Set (Cat. No. 562574/562725). FVS575V was also tested in mouse (data not shown).