Detection of the mouse acrosome reaction by acid phosphatase. Comparison with chlortetracycline and electron microscopy

The sperm acrosome is a uniquely regulated secretory vesicle containing sev eral hydrolase enzymes, including acid phosphatase (AP). The exocytotic eve nt that releases these enzymes, the acrosome reaction, is required for fert ilization in mammals. Different methods have been described in the scientif ic literature for detection of the acrosome reaction: double and triple sta ins, fluorescent-lectin stains, monoclonal antibodies against acrosomal ant igens (immunodetection techniques), Coomassie blue, differential interferen ce contrast or phase contrast, flow cytometry, and chlortetracycline (CTC). In contrast, only 1 method to detect AP released by live and reacted sperm has been described in the literature thus far. In this work we compare 2 c lassical methods, CTC and transmission electron microscopy (TEM), with the assay of AP released from the acrosome. AP released during the acrosome rea ction was measured in the culture medium. Enzyme remaining in nonreacted sp erm cells was released by Triton X-100 treatment. This enzyme-based methodo logy shows an increase of AP in the culture media after the acrosome reacti on and a corresponding decrease in the detergent-releasable enzyme. The AP assay thus permits the detection of the mouse acrosome reaction and compare s well with the CTC and TEM methods. This method is performed on the whole sperm population and so avoids the observer error that is inherent in light microscopic methods.