A new 96-well microtiter plate based adhesion assay was developed to measure weak cell adhesion. This assay is distinct from other adhesion assays by the procedure in which the nonadherent cells are removed. In most conventional adhesion assays, nonadherent cells are removed by aspiration followed by repeated washes. However, the shear force generated by such washing also detaches weakly adherent cells. In the minimal shear force adhesion assay (MSFA) described here, the removal of nonadherent cells is carried out by applying a gentle shear force in a fluid environment. In this procedure, adherent cells are not subjected to harsh and variable washing forces and are not exposed to surface tension caused by the removal of washing fluid between successive washes. Using the lymphoid cell lines XC1.5/51 and MPC11, the number of adherent cells determined by this new adhesion assay is three times higher than the conventional adhesion assay. This MSFA assay is simple, consistent, and easy to perform. With modifications for applying a defined shear force, this assay can be adopted to compare cell adhesion strength to various substrata.