YOPRO-1 PERMITS CYTOFLUOROMETRIC ANALYSIS OF PROGRAMMED CELL-DEATH (APOPTOSIS) WITHOUT INTERFERING WITH CELL VIABILITY

In the absence of cell permeabilization, the impermeant nuclear dye YO PRO-1 permits accurate analysis of apoptosis using cytofluorometry or fluorescent microscopy. Several immune cell populations were studied i ncluding dexamethasone-treated thymocytes, irradiated peripheral blood mononuclear cells and a growth factor-depleted tumor B cell line. Exc ellent correlation values were found with acridine orange using cytofl uorometry and with eosin-hematoxylin using optical microscopy. Under f luorescent microscopy, YOPRO-1-fluorescent cells demonstrate the morph ological features of cells undergoing apoptosis such as nuclear shrink age and fragmentation. An important characteristic of the dye that dif fers from all other nuclear dyes previously used for the detection of apoptosis is that it does not label living cells. Cell sorting after f low cytofluorometry analysis confirmed that only the apoptotic cell po pulation was labelled with YOPRO-1. Further studies showed that while incubation of living cells with Hoechst 33342 almost completely abroga ted the capacity of T cells to proliferate in response to several stim uli, YOPRO-1 had no inhibitory effect. This new simple, rapid and repr oducible use of the YOPRO-1 dye should prove useful in the analysis of apoptotic cells as well as for investigations of the functional prope rties of living cells in a culture containing apoptotic cells.