We have demonstrated earlier that the per sperm creatine-N-phosphotransferase (CK) activity was increased in oligospermic vs. normospermic men. The increased sperm CK activity is related to higher concentrations of cellular CK, which may indicate a defect of cytoplasmic extrusion during spermatogenesis. In the present work, we examined whether in spermatozoa, similar to muscle, there is a change in the synthesis of B-CK and M-CK isoforms during cellular differentation. In 109 normospermic and 50 oligospermic specimens (sperm concentrations 60.6 ± 3.7 vs. 8.8 ± 1.3 million sperm/ml; all values expressed as mean ± SEM), the relative concentrations of the M-CK isoform (M-CK / M-CK+B-CK) were 27.2% ± 2.1% vs. 6.7% ± 0.9% (P < 0.001). The per sperm CK activities showed comparable differences (0.21 ± 0.02 vs. 0.89 ± 0.1 CK IU/100 million sperm; P < 0.001) in the two groups, and there was a close correlation between per sperm CK activities and M-CK concentrations (R = 0.69, P < 0.001, N = 159). This indicates that the loss of cytoplasm and the commencement of M-CK isoform synthesis are related events during the last phase of spermatogenesis, also that the incidence of spermatozoa with incomplete cellular maturation is higher in oligospermic specimens. In characterizing the M-CK, we found that sperm (unlike muscle tissue) lack the MB hybrid of CK dimers. However, in the presence of muscle M-CK, the muscle-sperm MB-CK hybrid has formed. Thus in sperm and muscle the M-CK isoforms are structurally different, whereas the B-CKs are apparently homologous. We suggest that the relative concentrations of sperm M-CK isoform is a biochemical marker of cellular differentiation that may also predict the functional intetrity, such as fertilization potential, of spermatozoa.