Characterization of red blood cells agglutination based on stochastic collision electrochemical technique

Hemagglutination assays are important for blood group typing, alloantibodies screening and biomarkers detecting. Simple and sensitive approach for both detecting and quantifying hemagglutination is in need of development. In this research, the specific agglutination of red blood cells (RBCs) was characterized by stochastic collision events of RBCs on a Pt ultramicroelectrode (UME), extending the collision electrochemical technique to the biological reaction analysis. The collision and irreversible adsorption of RBCs blocked the oxidation flux on the UME and resulted in the "staircase" current drop in the chronoamperometry curve. The concentration, mean diameter and coverage ratio of RBCs were determined. Optical tracking of RBCs collision events verified the size and landing position dependence of RBCs to their collision step magnitudes. Adding specific antibodies to the RBCs, antibodies would connect RBCs together as a "bond" and form RBCs clusters. Such agglutination manifested itself by the decrease in collision frequency and the remarkable increase in the step current magnitude. Also, the agglutination extent could be roughly estimated from the largest step height or quantitatively determined from the decrease ratio of collision frequency. This work shows the reliability and sensitivity of collision electrochemical technique for hemagglutination analysis and builds the foundation for its further application on the blood-grouping sensors and immunoassays of infectious diseases.