Growth and expression of the 5F MGAT1^- CHO cell line expressing A244_N332-rgp120 in shake flask cultures.

<p>Cells were cultured under standard conditions until day 6 when 1 mM Sodium butyrate was added, and the temperature shifted to 32°C. Panels <b>A-C</b>: cells were fed with CHO Feed A and yeastolate as indicated, and harvested at day 13 (data from 3 shake flasks averaged). <b>(A)</b> Timecourse graph of viable cell densities (VCD) determined by trypan-blue exclusion on a BioRad T20 cell counter. <b>(B)</b> Timecourse of cell viabilities determined by trypan-blue exclusion. <b>(C)</b> Timecourse of A244_N332-rgp120 protein accumulation determined by ELISA. Panels <b>D-F</b> demonstrate optimization of protein expression (at >1g/L) by use of different feed additives. Five duplicate pairs of cultures were fed (as indicated) with CHO Feed C and either yeastolate (BD, Franklin Lakes NJ), cottonseed, wheat, pea hydrolsate (Friesland Camparia, Delhi, NY) or CD-hydrolysate (SAFC, Calsbad CA) at days 6, 8 and 10, and harvested at day 12 (data from each pair of shake flasks is averaged). <b>(D)</b> Timecourse graph of viable cell densities (VCD) determined by trypan-blue exclusion on a BioRad T20 cell counter. <b>(E)</b> Timecourse of cell viabilities determined by trypan-blue exclusion. <b>(F)</b> Timecourse of A244_N332-rgp120 protein accumulation determined by ELISA.