QUANTIFICATION OF CELL-MATRIX AND CELL-CELL ADHESION USING HORSERADISH-PEROXIDASE

A simple, universal, and rapid enzymatic method for the quantitative d etermination of cell adhesion in 96-well cell culture plates has been established. The assay is based on cellular steady-state endocytosis, which is used to label cells with horseradish peroxidase (HRP) prior t o adhesion. Subsequently, attached cells can be detected by a simple e nzymatic reaction, in which the accumulated HRP catalyzes dye formatio n from a colorless hydrogen donor, e.g., o-phenylenediamine, in the pr esence of hydrogen peroxide. As demonstrated with different cell lines and test systems, the method can be used to quantify cell-matrix as w ell as cell-cell interactions and allows a very sensitive quantificati on of adherent cells. The HRP label is nontoxic and does not affect th e adhesion properties of tested cell Lines; the quantity of dye formed is proportional to the number of adherent cells. Furthermore, the ass ay represents an alternative method to isotopic cell labeling, e.g., w ith Cr-51, which is usually used for quantifying cell-cell interaction s. (C) 1997 Academic Press, Inc.