Creatine kinase immunocytochemistry of human sperm-hemizona complexes: selective binding of sperm with mature creatine kinase-staining pattern

To examine the clinical significance of the increased sperm cytoplasmic content that is due to a fault of spermatogenesis, we have further studied the relationship between increased sperm creatine kinase (CK) concentrations and diminished fertilizing potential in men. In the present work, we used CK immunocytochemistry of human sperm-hemizona (HZ) complexes to examine whether the distribution of mature (clear heads), intermediate (sperm heads with light stippling), and immature (heads with heavy stippling or with solid CK staining) spermatozoa bound to the HZ would follow the incidence of these sperm in the samples tested, or if there is a preferential binding by the mature sperm.

Two pairs of HZ were exposed to washed semen and to their swim-up sperm fractions. The sperm and sperm-HZ complexes were treated with a CK antibody followed by horseradish peroxidase immunostaining, and the sperm were evaluated for maturity.

Men presenting for fertility evaluation were studied in two university-based andrology laboratories.

The binding of the HZ was selective for mature sperm as indicated by the incidence of intermediate and immature sperm in washed semen versus bound to the HZ (intermediate: 20.0% versus 1.4%; immature: 7.6% versus 0.5% [mean +/- SEM]) or in swim-up sperm fractions versus the HZ (intermediate: 18.7% versus 3.4%; immature: 2.5% versus 0.2%). The binding was almost exclusive to normal sperm (96.4% to 98.1%) whether the HZ were exposed to washed semen or swim-up fractions in spite of the five to ten times higher incidence of intermediate and immature sperm.

Mature sperm selectively bind to the zona. We suggest that spermatozoa with immature CK-staining patterns are deficient in the site(s) of oocyte recognition and binding.