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CE-based noncompetitive immunoassay for immunoglobulin G in bovine colostrum products
Jin Zhao1, Xiaojing Ding, Xinyu Wang
1Beijing Center for Disease Prevention and Control, Beijing, P. R. China.
Abstract:
A CE-based noncompetitive immunoassay for IgG in bovine colostrum products was established. FITC-labeled protein G (FITC-PrG) was tagged through noncovalent bindings to the Fc region of the mouse monoclonal antibovine IgG (Ab). The FITC-PrG, Ab, and IgG formed a sandwiched immunocomplex FITC-PrG-Ab-IgG under optimal incubation conditions. The immunocomplex was separated and analyzed by CZE with LIF detection in less than 2 min in an uncoated fused-silica capillary. Addition of PEG 20,000 (PEG 20M) in the running buffer significantly suppressed analyte adsorption and thus improved the reproducibility and the resolution. The precision of the method was 5.1% (n = 7). A linear relationship was established for the IgG concentration in the range of 1-5 mg/L with a linear correlation coefficient (r = 0.9917). The LOD was 0.1 mg/L (S/N = 3). The method was successfully applied for the determination of IgG in bovine colostrum products and satisfactory results were achieved.
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01:33Enzyme-Linked Immunosorbent Assay
There are many different types of ELISAs, but they all involve an antibody molecule whose constant region binds an enzyme, leaving the variable region free to bind its specific antigen. Enzyme-substrate reaction allows the antigen to be visualized or quantified.
01:15Western Blotting
The technique begins with separating proteins from the sample using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by protein transfer, immunoblotting, and finally, protein detection.