Enhancement of the Sensitivity of a Capillary Electrophoresis Immunoassay for Estradiol with Laser-induced Fluorescence Based on a Fluorescein-labeled Secondary Antibody

A competitive immunoassay for estradiol (E2) based on capillary electrophoresis was established. This method was based on the competitive reaction of complete antigen and E2 with a limited amount of monoclonal antibody, with fluorescein isothiocyanate (FITC)-labeled secondary antibody as a fluorescence probe. The addition of the thermally reversible hydrogel, poly-N-isopropylacrylamide (pNIPA) in the buffer as a replaceable packing material improved reproducibility and resolution of the method. This capillary electrophoresis immunoassay with laser-induced fluorescence can be applied to determine E2 with good precision at concentrations as low as 9 pg/mL. Details of typical separations of immunological complex and free reactants are presented.