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Immunomagnetic Isolation of Fetal Rat Gonocytes
Author(s) -
Ham R.,
Pelt A.M.M.,
Miguel M.P.,
Kooten P.J.S.,
Walther N.,
DisselEmiliani F.M.F
Publication year - 1997
Publication title -
american journal of reproductive immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.071
H-Index - 97
eISSN - 1600-0897
pISSN - 1046-7408
DOI - 10.1111/j.1600-0897.1997.tb00274.x
Subject(s) - gonocyte , collagenase , andrology , biology , fetus , microbiology and biotechnology , monoclonal antibody , trypsin , antigen , embryo , antibody , chemistry , immunology , biochemistry , enzyme , pregnancy , medicine , genetics
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 10 3 ; 18 days pc) or 1.2 ± 0.25 (x 10 4 ; 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.