Optimization of Key Factors in Serum Free Medium for Production of Human Recombinant GM-CSF Using Response Surface Methodology.

Abstract

Researchers add serum to a classical medium at concentrations of 5 to 10% (v/v) to grow cells in-vitro culture media. Unfortunately, serum is a poorly defined culture medium component as its composition can vary considerably while serum-free cell culture media are an excellent alternative to standard serum-containing media and offer several major advantages. Advantages of using serum-free media include a lower risk of infectious agents, lower risk of interfering components, less contaminant, avoids ethical issues. According to previous studies insulin, selenium, transferrin and glucose are important component of serum that affect cell growth. In the present study, we optimized amount of these factors in order to serum free culture medium fabrication. Response surface methodology (RSM) was employed for optimization of key factors in serum free medium to enhance recombinant human GM-CSF (rhGM-CSF) production in CHO cell line. Four important process parameters including insulin concentration (0-2 g/L), transferrin concentration (0-1 g/L), selenium concentration (0-0.001 g/L) and glucose concentration (0-5 g/L) were optimized to obtain the best response of rhGM-CSF production using the statistical Box–Behnken design. The experimental data obtained were analyzed by analysis of variance (ANOVA) and fitted to a second-order polynomial equation using multiple regression analysis. Numerical optimization applying desirability function was used to identify the optimum conditions for maximum production of rhGM-CSF. The optimum conditions were found to be insulin concentration = 1.1 g/L, transferrin concentration = 0.545 g/L, selenium concentration = 0.000724 g/L and glucose = 1. 4 g/L. Maximum rhGM-CSF production was found to be 3.5 g/L.