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Simply Mixing Poly Protein G with Detection Antibodies Enhances the Detection Limit and Sensitivity of Immunoassays

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journal contribution
posted on 30.05.2019, 00:00 by Yi-Jou Chen, Michael Chen, Tian-Lu Cheng, Steve R. Roffler, Shyr-Yi Lin, Hui-Lan Hsu, Chang-Hung Wang, Che-Yi Chen, An-Pei Kao, Jing-Jy Cheng, Kuo-Hsiang Chuang
An insufficient amount of detection antibodies bound to their antigens usually limits the sensitivity of immunoassays. Here, we describe a simple method to improve the detection limit and sensitivity of various immunoassays by mixing detection antibodies with a soluble poly protein G (named 8pG). 8pG was developed by fusing eight repeated fragment crystallizable (Fc) binding domains of streptococcal protein G to a linear polymer. Simply mixing detection antibodies with 8pG to form an antibody/8pG complex largely increased the accumulation of detection antibody to target molecules, which dramatically enhanced the sensitivity in direct ELISA, sandwich ELISA, Western blot, and flow cytometry systems, separately. The detection limit of Western blot for low-abundance PEGylated interferon (Pegasys) and recombinant human CTLA4 (rhCTLA4) improved by at least 13-fold and 31-fold, respectively, upon mixing detection antibodies with 8pG. Moreover, the nanoscale size of the antibody/8pG complex did not influence the granularity and dimension of target cells in the flow cytometry system. Collectively, we provide a quick and easy-to-operate method to make various immunoassays to sensitively detect low-abundance target molecules by just mixing their detection antibodies with 8pG.