The Propagation of Mammalian Cells in a 20 - Liter

The propagation of mammalian cells in stainless steel fermentors of a type used in antibiotic fermentations represents a marked departure from the techniques employed in classical tissue culture studies. Since the pioneer work of Harrison (1907), in which he demonstrated the outgrowth of cells from nerve tissue maintained in vitro, many techniques for the in vitro culture of cells have been developed. An older method of cell culture, which is still used, coinsists of imbedding small fragments of excised tissue in a drop of nutrient medium on the glass surface of a test tube or flask. The medium generally includes embryo extracts and blood plasma which form a coagulum about the tissue mass. The imbedded tissue may be bathed in a nutrient solution, composed of serum, carbohydrate, amino acids, or protein hydrolyzates, and salts. The small amount of tissue produced as well as the nature of environmental conditions make application of this system to viral, nutritional, and physiological studies most difficult (Parker, 1950). Aniother technique, the cultivation of cells as a monolayer on glass surfaces, was described by Carrel and EIbeling (1922). Today the culture of cells as monolayers in flasks and bottles is the most widely used method (Hanks et al., 1955). The preparation of cells for monolayer culture requires their dispersion from clumps and tissue masses by mechanical means or by the use of agents such as trypsin and Versene. The dispersed cells are suspended in a nutrient medium and allowed to settle onto the bottom of the glast vessel where they adhere to the glass surface and multiply. This method makes possible investigations concerned with the nutrition of the cells as well as the kinetics of virus-host cell reactions. However, the amount of cells which can be produced is necessarily limited and (1uantitation of cell responses to imposed conditions lacks precision. Recent investigations have established the feasi-