Detection and kinetics of the normal acrosome reaction of mouse sperm

Abstract A procedure is described for quantifying the percentage of normal acrosome reactions in fixed mouse sperm. This technique, which is a modification of the triple stain technique (TST) devised for human sperm [Talbot and Chacon, 1981a, b], utilizes trypan blue exclusion to identify the percentage of dead sperm in culture and subsequent staining of fixed sperm to differentiate the acrosome. Four classes of mouse sperm can be distinguished with the TST: (1) “live,” unreacted sperm, (2) “live,” acrosome‐reacted sperm (normal reactions), (3) dead, unreacted sperm, and (4) dead, acrosome‐reacted sperm (degenerative acrosome reactions). We have used this technique to study the time course of the normal acrosome reaction of mouse sperm in vitro. Our results with TST show that a low percentage of mouse sperm undergo normal acrosome reactions during the first 30 min of incubation; a maximum level of reactions is reached by 2 hr. Calcium was required in the medium for occurrence of normal reactions.