Critical factors in establishing monolayer cultures of normal human placental cells in serum-free medium.

Comparative studies have been performed to determine the best method for preparing monolayer cultures of normal human term trophoblast cells. The use of 0.25% trypsin-10 U/ml DNAse I provided the highest cell viability and greatest hormone production, but was critically dependent on the trypsin lot used. Cell function in culture was not improved with various substrata, nor did other factors (medium type, pH, red blood cell removal) affect the results. Optimization of dispersal and culture conditions permitted the trophoblast cells to survive with intact hormone secretion and response to secretagogues under serum-free conditions. These studies thus define the best methodology for establishing trophoblast cells in monolayer cultures.