Isolation and preliminary characterization of pig primordial follicles

Summary. An enzymic method for recovering primordial follicles from the pig ovary consists of incubating cortical slices for 2 h with 0\m=.\025%collagenase 1A. An average of 185 000 or 419 000 primordial follicles per ovary were recovered from ovaries collected in Cambridge and Kansas, respectively. Following a discontinuous Percoll gradient, primordial follicles can be separated from contaminating somatic cells by mouth pipette or a micromanipulator to collect 100\p=n-\1500follicles but for large scale recovery of approximately 30 000 follicles flow cytometry is recommended. Two types of pri-mordial follicles can be distinguished by electron microscopy: peripheral clusters of small oocytes with an incomplete investment of pregranulosa cells and a deeper region of individual oocytes surrounded by a complete layer of pregranulosa cells. The viabi-lity of the purified primordial follicles is attested by their ability to synthesize proteins for at least 12 h after incubation with [35S]methionine. Moreover, the primordial fol-licles showed several polypeptide bands in common with mature oocytes especially with Mr of 60 000\p=n-\90000 but with considerable differences from somatic cells