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 (vc) 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 vc or vice versa. In this work, the agglutination strength was derived from the vc 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 vc values of different samples were correlated with their agglutination strengths at a given WSS and antibody surface density. The vc decreased as the agglutination strength increased, which can be considered as a linear regression. The coefficient of variation of the calculated vc decreased to 0.1 as vc 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.

Blood Group Testing

“…Generally, RBC antigen-specific IgM or IgG antibodies immobilized on the surface of SPR can be used to identify blood groups (71,72). The intensity of agglutination depends on RBC adhesion and wall shear stress (73). The adhered RBCs move at different average cell velocities under the shear flow, which produces resistance.…”

“…Higher average cell velocity decreases the resistance of antigen and antibody binding and lowers agglutination intensity. The antigen density of A 1 and B RBCs from that of A 1 B RBCs and can be resolved using this approach (73).…”

Application of Long-Range Surface Plasmon Resonance for ABO Blood Typing

“…Houngkamhang et al [27] reported ABO blood typing via SPR imaging which observed interaction between immobilized Anti-A and Anti-B antibodies array and A and B antigens on RBCs surface. The interaction of antigens around lower RBCs membrane surface and immobilized Anti-A and Anti-B antibodies array was applied to obtain the agglutination strength of the RBCs and immobilized antibody [28]. The SPR imaging, which utilized the shear force generated within the flow cell, was used to measure the rolling speed of red blood cell [28].…”

“…The interaction of antigens around lower RBCs membrane surface and immobilized Anti-A and Anti-B antibodies array was applied to obtain the agglutination strength of the RBCs and immobilized antibody [28]. The SPR imaging, which utilized the shear force generated within the flow cell, was used to measure the rolling speed of red blood cell [28]. The technique can only extract the information at the interface because the limit of the evanescence field makes it impossible to study in whole cell.…”