Noncompetitive immunoassays using protein G affinity capillary chromatography and capillary electrophoresis with laser-induced fluorescence detection

Abstract

A new and simple approach to perform immunoassay using protein G affinity capillary chromatography and laser-induced fluorescence detection was described. A noncompetitive assay for monoclonal anti-bovine serum albumin (BSA) was used to test the performance of the system. Fluorescein isothiocyanate labeled BSA was used as a tracer to determine anti-BSA in p M level. Capillaries with inner diameter of 150 μm were packed with recombinant protein G-bound perfusive support. The packed capillary was used to capture the immunocomplexes, which were subsequently desorbed by 100 m M glycine (pH 9.0). Open tube capillary electrophoresis-based immunoassay (CEIA) for anti-BSA was also performed. Using standard samples, calibration curves for anti-BSA was established in both assays. Compared with CEIA, this system improved the concentration sensitivity for about 100-fold due to the pre-concentration of immunocomplex in the protein G column, while the mass sensitivity was similar in the two methods.