Thus our flow cytometric approach, which uses minimally low concentrations of zona material, which obviates the need for pretreatment or washing of the cells (thereby minimizing cell damage), and which enables analysis of some 10 000 cells in less than 15 sec, would seem to provide a sensitive and quantitative evaluation of binding of zona pellucida components to the surface of intact spermatozoa. By including the viability probe PI in our media, we were able to distinguish between live and dead sperm in our suspensions, and thereby, crucially, to focus on the live cells; only the latter could be expected to participate in the fertilization process. birth control pills
Enhanced sperm death during incubation in bicarbonate-containing Tyrode’s based media (i.e., under fertilizing conditions) has been reported previously . Indeed, it has been proposed that capacitation (necessarily induced by IVF conditions) is an ongoing destabilization process that inevitably and eventually leads to cell death . At present, the most likely explanation for the two levels of zona binding that developed in the dead cell population (upper right quadrant in Fig. 2b) is that these represented terminally degenerated cells derived from each of the two live subpopulations.