Extracellular Matrix Interferes with Colorimetric Estimation of Cell Number

Colorimetric methods for enumerating cultured cells have become very popular due to their applicability to a 96-well microplate format and the simplicity of their methodology. They have largely replaced direct counting methods, which are tedious to do and require relatively large cell numbers. Colorimetric assays fall into two categories; those that measure the activity of a particular cellular enzyme and those that stain cellular components. Use of the former requires that the individual cellular metabolic activity is similar over the range of cell densities and experimental conditions studied (Connolly et al., 1986; Hansen et al., 1989). The staining techniques require a low background from surface coatings or extracellular matrix produced by the cells themselves (Skehan et al., 1990; Muir et al., 1993; Schultz et al., 1994). We have tested representatives of each type of method to estimate the densities of cultured human umbilical artery endothelial (HUAEC) and human adult mammary artery vascular smooth muscle cells (VSMC).