Evaluation of agglutination strength by a flow-induced cell movement assay based surface plasmon resonance (SPR) technique

Abstract

A flow-induced cell movement assay combined with a surface plasmon resonance (SPR) technique was developed to quantify the agglutination strength, derived from the standard tube-agglutination test. Red blood cells (RBCs), based on the ABO blood group system, were specifically captured by anti-A and/or anti-B antibodies immobilized on a sensor surface. The agglutination strength corresponds to the amount of antigen–antibody interactions or the strength of RBC adhesion. Under a shear flow, the adherent RBCs were forced to move out of the region of interest with different average cell velocities (v_c) depending upon the adhesion strength and wall shear stress (WSS). That is, a higher adhesion strength (higher agglutination strength) or lower WSS represents a lower v_c or vice versa. In this work, the agglutination strength was derived from the v_c that was calculated from the time derivative of the relative SPR signal by using a simple model of cell movement response, whose validity was verified. The v_c values of different samples were correlated with their agglutination strengths at a given WSS and antibody surface density. The v_c decreased as the agglutination strength increased, which can be considered as a linear regression. The coefficient of variation of the calculated v_c decreased to 0.1 as v_c increased to 30 μm min⁻¹. The sensitivity of this assay can be controlled by optimizing the antibody surface density or the WSS. This assay has the capability to resolve the antigen density of A₁ and B RBCs from that of A₁B RBCs.