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Basic Techniques in Mammalian Cell UNIT 1.1 Tissue Culture Mary C. Phelan1 1Molecular Pathology Laboratory Network, Maryville, Tennessee ABSTRACT Cultured mammalian cells are used extensively in cell biology studies. It requires a number of special skills in order to be able to preserve the structure, function, behavior, and biology of the cells in culture. This unit describes the basic skills required to maintain and preserve cell cultures: maintaining aseptic technique, preparing media with the appropriate characteristics, passaging, freezing and storage, recovering frozen stocks, C and counting viable cells. Curr. Protoc. Cell Biol. 36:1.1.1-1.1.18. 2007 by John Wiley Sons, Inc. Keywords: mammalian cells tissue culture aseptic technique medium passaging cells freezing cells Tissue culture technology has found wide application in the field of cell biology. Cell cultures are utilized in cytogenetic, biochemical, and molecular laboratories for research as well as diagnostic studies. In most cases, cells or tissues must be grown in culture for days or weeks to obtain sufficient numbers of cells for analysis. Maintenance of cells in long-term culture requires strict adherence to aseptic technique to avoid contamination and potential loss of valuable cell lines (see UNIT 1.3). An important factor influencing the growth of cells in culture is the choice of tissue culture medium. Many different recipes for tissue culture media are available and each laboratory must determine which medium best suits its needs. Individual laboratories may elect to use commercially prepared medium or prepare their own. Commercially available medium can be obtained as a sterile and ready-to-use liquid, in a concentrated liquid form, or in