A method for measuring specific IgE in sera by direct ELISA without interference by IgG competition or IgG autoantibodies to IgE

Background In measuring specific IgE levels in sera by direct ELISA, compet ition with coexisting IgG often impedes an exact IgE determination; additio nally, IgG autoantibodies to IgE (IgG-IgE) in sera affect the assay. in thi s paper, we attempt to determine accurate specific IgE levels by selective removal of IgG with a protein G-immobilized gel (PG) and by acid treatment of the PG to compensate for the unintended removal of IgE, probably due to the PG binding IgG-IgE, Methods: IgG in sera was removed using PG at pH 7.0 . Then, the PG was treated with citrate buffer at pH 3.0 for 5 min to liber ate IgE from IgG-IgE complexes, after IgG-binding sites on the PG were satu rated with bovine IgG, since PG came to bind IgE at acidic pi-is. IgE level s were then measured by ELISA. Results: The PG treatment of sera removed th e effect of inhibitory competition by coexisting IgG, especially at higher concentrations of sera, to improve specific IgE detection by direct ELISA, However, PG treatment alone sometimes reduced IgE levels (39% of sera teste d), even though PG does not bind IgE at pH 7.0, which indicated the presenc e of IgG-IgE complexes. The reduction in IgE returned almost to their origi nal levels in the sera by acid treatment of the PG, By combining the PG tre atment with acid treatment, specific IgE measurement in sera was improved s ignificantly (p < 0.01, Wilcoxon signed rank test). Conclusion: Measurement of specific IgE in sera by direct ELISA was improved by using the PG and a cid treatment technique. Copyright (C) 2000 S. Karger AG,Basel.