AbstractCulture preparations vary greatly in complexity, ranging from single isolated cells to three‐dimensional histotypic cell structures. Besides cultures obtained directly from animal tissues (primary cultures), permanent cultures of continuously dividing (immortalized) cells have been established. This unit highlights both the advantages and disadvantages of a number in vitro approaches including primary cultures, continuous (permanent or immortal) cell lines, and contrasts culture techniques including suspension, attached (monolayer), and three‐dimensional (aggregate) cultures in addition to explants. The basics of culture maintenance and propagation are covered (i.e., tissue dispersion, cell separation and purification, and passaging), and information is provided on the critical aspects of culturing cells, such as pH of the media, osmolality, humidity, and cell density. Also included is a troubleshooting section on how to cope with problems of contamination by bacteria, mycoplasma, toxic chemicals or foreign cell types.Culture preparations vary greatly in complexity, ranging from single isolated cells to three‐dimensional histotypic cell structure.