Long-term culture of bovine trophoblastic cells

Abstract

Procedures for the isolation and cultivation of bovine cotyledonary trophoblastic cells from early second trimester placentas are described. Collagenase digestion yielded a single cell suspension of both uninucleate and binucleate cells with minimal contamination with other cell types. Optimal growth conditions were obtained through supplementation of medium with epidermal growth factor, insulin, transferrin, and selenium. Trophoblastic cells survived multiple passages, cryopreservation, and serum deprivation and have been maintained in culture for more than 14 mo. Two cell types were identified by phase microscopy: uninucleated trophoblastic cells and trophoblastic giant cells, which were predominantly binucleate cells. Binucleate cells were present in small numbers through all passages. This procedure provides a reliable method to obtain trophoblastic cell lines for studies of trophoblastic cell physiology and susceptibility to infectious and toxic agents.