Kinetic study of the initial stages of agglutination process with scanning flow cytometer

The main disadvantage in ordinary agglutination immunoassay is difficulties to distinguish between specific and non-specific particle aggregation. We proposed to use Scanning Flow Cytometry to the kinetic study of the initial stages of agglutination process. The main advantage of the Scanning Flow Cytometry is a possibility to measure angular dependency of the light scattered by a single particle, an indicatrix. The most promising field for application of the indicatrix technology is a characterization of non-spherical particles. Validity of proposed method was verified by simultaneous measurements of the light scattering and fluorescence signal. We used Wentzel-Kramers-Brillouin approximation to simulate light scattering from two glued spheres and to explain the results obtained from measured indicatrices. To show an applicability of the proposed technique, the kinetic experiments were performed on latex particle covered with BSA (diameter 1.8 μm). Kinetics of dimer fraction growth initiated by mixing BSA-covered latex particles with anti-BSA immuoglobulins IgG was studied. In order to evaluate kinetic rate constant simple kinetic model involved only dimer growth reaction was applied for data treatment. Two kinetic rate constant for dimer fraction growth k_B=2.88•10^-12 cm³s^-1 and k_A=0.85•10^-12 cm³s^-1 were evaluated for two samples with the same origin but with different prehistory.