Reducing the Requirement for Serum Supplements in High Yield Microcarrier Cell Culture

Abstract

Various methods of reducing the requirement and cost of serum supplements for the growth of MRC-5 and Vero cells on Cytodex^R microcarriers were examined. The base medium was DME/Medium 199 (50/50) and a supplement of 10% (v/v) foetal calf serum (FCS) was the control. Greater economy of serum could be achieved by selecting the types and concentration of serum on the basis of whether the serum was to support the attachment and growth of cells at low densities or at later stages in the culture cycle. The concept of using a fixed concentration of serum throughout the culture cycle is questioned.

Low-serum and serum-free media could support the growth of cells on Cytodex 1. Medium supplemented with 0.5% FCS, bovine serum albumin (BSA, 2 mg/ml) and either EGF (10 ng/ml), or insulin (1 μg/ml) and transferrin (25 μg/ml), proved to be satisfactory for stirred cultures and supported cell proliferation to nearly the same extent as the control. Various serum-free media were tested and maximum yields were obtained when the base medium was supplemented with fibronectin (2 μg/ml), BSA (2 mg/ml), insulin (1 μg/ml), transferrin (25 μg/ml), putrescine (100 μM), fetuin (1 mg/ml) and EGF (10 ng/ml). Replacement of fetuin by dexamethasone (50 ng/ml) and trace metals (Mo, Cd, Se) resulted in a small reduction in cell proliferation. Cell yield was approx. 40–50% of that obtained with 10% FCS. A possible protective effect of larger molecules in low-serum and serum-free media may be important for reproducible results with stirred microcarrier cultures.