Magnetic cell sorting is a fast and effective method of enriching viable spermatogonia from djungarian hamster, mouse, and marmoset monkey testes

Germ cell transplantation, which offers promising new approaches for resear ch and clinical applications, has focused interest on spermatogonia. This p aper describes a procedure that permits the isolation of large quantities o f viable spermatogonia. The immunomagnetic isolation procedure was applied to testicular cell suspensions from photoinhibited and photostimulated Djun garian hamsters, mice, and marmoset monkeys. The cells were incubated with a polyclonal rabbit anti-c-kit IgG, binding of which was characterized by i mmunohistochemical staining. For magnetic labeling, a secondary anti-rabbit Ige conjugated to ferromagnetic microbeads was used. Separation columns al lowed the retention of magnetically labeled cells within the matrix. The ma gnetic fractions were eluted after removal of the column from the magnetic field. All fractions were analyzed for cellular morphology and by flow cyto metry. The final enrichment of c-kit-positive cells in the magnetic fractio n using fully active testes was in the range of 25-55% with a viability rat e of so-sox. The magnetic fractions of all three species were characterized by high numbers of diploid cells. Cytological analysis revealed a strong e nrichment of spermatogonia. No haploid cells were retained in the magnetic fraction. In comparison to conventional procedures, magnetic cell separatio n is an efficient and fast approach for isolation of spermatogonia.