Clonal agar culture of normal primary explanted bovine granulosa cells.

When seeded at low cell densities (500 to 2500 cells per dish), normal primary explanted bovine granulosa cells grow in a double layer semi-solid agar culture system as anchorage-independent clones composed of well differentiated cells with vacuolated granular cytoplasm with a cloning efficiency of between 2% and 4%. The addition of growth promoting factors to the culture medium such as epidermal growth factor (EGF), fibroblast growth factor (PGF) or rat erythrocyte suspensions resulted in a 5 to 6-fold increase in colony numbers and this was accompanied by an increase in colony size. Cells from both small (3--7 mm diameter) and large (less than 1 cm diameter) ovarian follicles exhibited similar patterns of responsiveness to growth factors but the cloning efficiency of cells from large follicles was approximately double that of cells from small follicles.