Validation of Flow Cytometry Analysis in the Objective Assessment of Spermatogenesis: Comparison to the Quantitative Testicular Biopsy

Abstract

Objective determination of spermatogenesis has been accomplished by quantitative testicular biopsy, which, although laborious, has served as the standard for spermatogenic assessment. Aspiration deoxyribonucleic acid (DNA) flow cytometry of the testis, however, has simplified this determination, and has correlated with indirect hormonal parameters of spermatogenesis and qualitative observations of the seminiferous epithelium. Nevertheless, this important modality has yet to be validated against quantitative micrometry of the testis. To determine this correlation we submitted 29 incisional testicular biopsies for simultaneous quantitative analysis and DNA flow cytometry. Micrometric parameters included the mean tubular wall thickness, and the mean tubular concentration of late spermatids and Sertoli cells. The percentage of haploid, diploid and tetraploid cells was determined for each patient. For the entire patient population a statistically significant correlation was observed between the percentage of haploid cells and the tubular concentration of late spermatids (r = 0.784, p <0.0005) as well as the mean tubular spermatid-to-Sertoli cell ratio (r = 0.824, p <0.0005). A similar correlation was noted for various etiological subsets of patients: spinal cord injury (r = 0.809, p <0.002), genital tract obstruction (r = 0.705, p <0.02) and miscellaneous diagnoses (r = 0.828, p <0.02). For the group with testicular failure quantitative micrometry and flow cytometry demonstrated severe impairment in all patients although a statistically significant correlation could not be shown because of the small range of values. DNA flow cytometry analysis correlates strongly with the current standard of quantitative spermatogenic assessment and, therefore, it may be validated as a simplified and highly objective method of determining spermatogenesis.