Cell Adhesion Assays

Standard adhesion assays measure cell binding either to immobilized ligands or to cell monolayers in flat-well microtiter plates under static conditions. Typically, these test systems require several washing steps to separate adherent from nonadherent cells. Here, we describe an adhesion assay which avoids these washing steps by employing V-bottom 96-well plates. In this assay, fluorescently labeled leukocytes are allowed to adhere to V-well plates coated with soluble ligand for a fixed time. Thereafter, centrifugal force is applied to separate adherent cells from nonadherent cells. Nonadherent cells accumulate in the nadir of the V-shaped wells and are quantified using a fluorometer with a narrow aperture. This simple and reproducible method has been validated with different classes of adhesion molecule families (selectins and integrins) and is adaptable to several other adhesive interactions. The assay format is suitable for screening applications and may also be used for diagnostic testing. The receptor/ligand interaction chosen as an example to describe the assay methodology is the interaction between the integrin lymphocyte function-associated molecule-1 (LFA-1, αLβ2) and intercellular adhesion molecule-1 (ICAM-1).