ANALYSIS OF IMMUNOGLOBULIN-G USING A CAPILLARY ELECTROPHORETIC AFFINITY ASSAY WITH PROTEIN-A AND LASER-INDUCED FLUORESCENCE DETECTION

A method for the rapid and sensitive determination of immunoglobulin G (IgG) in cultivation media by an affinity assay using capillary elect rophoresis is presented. For that purpose we evaluated protein A conju gated with a fluorescent dye such as fluorescein diisocyanate or dichl orotriazinyl-aminofluorescein as an affinity ligand. The ligand formed a fluorescing complex with immunoglobulin G in the sample and rapid s eparation from excess protein A was performed by capillary zone electr ophoresis. However, only partial resolution of the zones was achieved when protein A as a whole molecule was utilized. In contrast, baseline resolution of the zones was obtained when recombinant fragments of pr otein A were used as affinity ligands. Immunoglobulin concentrations i n the range of two orders of magnitude were determined. Due to the spe cifity of protein A for immunoglobulin G, analysis can be carried out even in the presence of high concentrations of other components and in cultivation media. Thus, the capillary electrophoretic affinity assay was successfully applied to monitor monoclonal antibodies in a cultiv ation process.