Immunomagnetic isolation of fetal rat gonocytes.
OBJECTIVE
An efficient method to obtain highly enriched populations of viable gonocytes from rat embryos at Day 18 and Day 20 postcoitum (pc) is described.


METHODS
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, 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 (x10(3); 18 days pc) or 1.2 +/- 0.25 (x10(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.


CONCLUSIONS
The cells were successfully used for culture studies and as starting material for the investigation of gene expression.
van den Ham
R
R
Department of Functional Morphology, Veterinary School, Utrecht University, The Netherlands.
van Pelt
A M
AM
de Miguel
M P
MP
van Kooten
P J
PJ
Walther
N
N
van Dissel-Emiliani
F M
FM
D000818:Animals; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D004622:Embryo, Mammalian; D005260:Female; D007075:Immunoglobulin M; D018189:Immunomagnetic Separation / Q000379:methods*; D008297:Male; D011247:Pregnancy; D012333:RNA, Messenger / Q000302:isolation & purification; D051381:Rats; D017208:Rats, Wistar; D013737:Testis / Q000737:chemistry / Q000166:cytology*
D000911:Antibodies, Monoclonal; D007075:Immunoglobulin M; D012333:RNA, Messenger