Preparation of enriched mouse syncitia-free pachytene spermatocyte cell suspensions

Spermatogenesis comprises a complex succession of steps of mitosis, meiosis, and differentiation, starting with the commitment of diploid spermatogonial stem cells to differentiate and ending with the formation of haploid spermatozoa. Rodent models have been routinely used to study germ cell development and reproductive toxicology, since they present many similarities with their human counterpart while offering the advantage of recapitulating within a few weeks a process that normally takes years to occur. This article describes a method to isolate subpopulations of adult germ cells, more specifically pachytene spermatocytes, using two successive gradients, without using an elutriation centrifuge, a specialized device not available in many laboratories. Moreover, the method was designed to isolate enriched pachytene spermatocytes preparations devoid of contaminating syncitia, often formed in response to toxicants or environmental insults.