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Additional resources for Animal Cell Culture: A Practical Approach 3rd Edition
8 is usually inhibitory to cell growth. Factors affecting the pH stability of the medium are buffer capacity and type, headspace, and glucose concentration. The normal buffer system in tissue culture media is the carbon dioxide bicarbonate system analogous to that in blood. This is a weak buffer system, in that it has a pKa well below the physiological optimum. It also requires the addition of carbon dioxide to the headspace above the medium to prevent the 24 SCALING-UP OF ANIMAL CELL CULTURES loss of carbon dioxide and an increase in hydroxyl ions.
53. 54. 55. 18 Eagle, H. (1959). Science, 130, 432. Dulbecco, R. and Freeman, G. (1959). Virology, 8, 396. Ham, R. G. (1965). Proc. Natl. Acad. Sci. USA, 53, 288. Barnes, D. and Sato, G. (1980). Anal. , 102, 255. Leibovitz, A. (1963). Am. J. , 78, 173. Peehl, D. M. and Ham, R. G. (1980). In Vitro, 16, 526. Stacey, G. , Masters, J. R. , Hay, R. , Drexler, H. , MacLeod, R. A. F. and Freshney, R. I. (2000). Cell contamination leads to inaccurate data: we must take action now. Nature, 356. K. 1 Introduction Small scale culture of cells in flasks of up to 1 litre volume (175 cm2 surface area) is the best means of establishing new cell lines in culture, for studying cell morphology and for comparing the effects of agents on growth and metabolism.
LDH is measured in cell-free medium at 30°C by following the oxidation of NADH by the change in absorbance at 340 nm. The reaction is initiated by the addition of pyruvate (2). One unit of activity is defined as 1 umol/min NADH consumed. LDH is released by dead/dying cells and is therefore a quantitative measurement of loss of cell viability. To measure viable cells a reverse assay can be performed by controlled lysis of the cells and measuring the increase in LDH. 1 Culture vessel and growth surfaces The standard non-disposable material for growth of animal cells is glass, although this is replaced by stainless steel in larger cultures.