A comparison of three methods for detecting the acrosome reaction in human spermatozoa

This study was designed to compare three different fluorescent probes to assay the acrosome reaction in human spermatozoa: chlortetracycline (CTC), mannosylated bovine serum albumin (BSA) labelled with fluorescein (MAF), and quinacrine (QN). Normal human sperm ejaculates were washed and allowed to swim up for 30-60 min. Samples were examined under epifluorescence for the percentage of the acrosome reacted spermatozoa, as detected by the three probes. There was no significant differences between samples of fresh, uncapacitated spermatozoa evaluated with CTC, MAF or QN; all gave < 10% reacted. Following capacitation for 3 h, the percentage of spontaneously reacted spermatozoa was higher than in fresh spermatozoa; CTC and MAF gave the same percentage (12%), while QN indicated a higher percentage (18%) of reacted spermatozoa (P < 0.001). Following exposure to ionophore A23187 at 1 h, the percentage of acrosome reactions increased to a mean of 31% as detected with CTC or MAF; the mean percentage (45%) was significantly higher with QN (P < 0.0001). Further incubation up to 2 h with A23187 did not change these percentages. These results suggest that the QN probe detects the onset stage of the acrosome reaction, whereas the CTC and MAF probes detect the later stages in which the acrosomal cap is lost. Use of the two types of probe provides a means for finer resolution of the time course of the acrosome reaction in the human spermatozoa.