A novel image cytometric method for quantitation of immunohistochemical staining of cytoplasmic antigens

A novel method was developed to evaluate the acrosomal status of mammalian spermatozoa. The method is based on the ability of the lectin Pisum sativum agglutinin (PSA) to bind specifically to glycoproteins of the acrosomal matrix released during the acrosome reaction. The amount of released acrosomal content is proportional to the fraction of spermatozoa that underwent acrosome reaction. The released glycoproteins present in the supernatant separated from the cells were detected via an ELISA-like assay. The authors suggest that one of these glycoproteins might be the acrosin as identified by anti-acrosin antibodies, using Western blot analysis. The new method (demonstrated here with ram and bull spermatozoa) correlates well with the results obtained by conventional methods. Its advantages are simplicity, objectiveness, rapidity, and low cost. In addition, many samples can be processed in parallel. The method can be used in experimental as well as clinical applications.