The glucose, insulin and glutamine requirements of suspension cultures of HeLa cells in a defined culture medium.

The serum supplement in a defined medium for the growth of HeLa cells could be replaced by protamine-zinc-insulin (02 u./ml). Insulin (0-4 u./ml) replaced the growth-stimulatory properties of protamine-zinc-insulin, whilst protamine sulphate (5 /tg/ml) was found to be toxic to the cells. The addition of insulin to cultures depleted of insulin increased both cell growth rates and maximum cell populations. In the denned medium, HeLa cells could only utilize glutamate when a small amount of glutamine was included. Glucose, at a level of 2 mg/ml, was shown to limit maximum cell populations. The growth yield from glucose was 295 fig cell dry weight/mg glucose. When the medium glucose concentration was increased to 4 mg/ml, HeLa cell populations in excess of 16 x 10 cells (i.e. 640 fig dry weight)/ml were routinely achieved in the denned medium supplemented with insulin. Growth is then limited by the amino acid supply. Increasing the amino acid concentration of the medium by 50 % raised the maximum cell population to 235 x io cells (i.e. 940 fig dry weight)/ml.