Immunomagnetic Isolation of Fetal Rat Gonocytes

Authors


Department of Functional Morphology, Veterinary School, Utrecht University, PO Box 80.157, 3508 TD Utrecht, The Netherlands.

Abstract

PROBLEM: An efficient method to obtain highly enriched populations of viable gonocytes from rat embryos at Day 18 and Day 20 postcoïtum (pc) is described.

METHOD: Single-cell suspensions with high cell yield were obtained by a collagenase/ trypsin digestion of the decapsulated testis. The gonocytes were purified by a direct immunoseparation technique,1,3 using magnetizable beads coated with rat anti-mouse immunoglobulin M (IgM) and a monoclonal antibody 4B6.3E10, which specifically reacted with a differentiation antigen on the fetal germ cells.

RESULTS: Populations of 8.3 ± 2.7 (x 103; 18 days pc) or 1.2 ± 0.25 (x 104; 20 days pc) viable gonocytes per testis with purities of 91 ± 6.5% and 92 ± 4.3%, respectively, as determined by Nomarski microscopy were obtained.

CONCLUSION: The cells were successfully used for culture studies and as starting material for the investigation of gene expression.

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