The Construction Of Antibody And Horseradish Peroxidase Polymer

Objects:We constructed two antibody and horseradish peroxidase polymers by chemical method. One is IgG-PLL-HRP whose skeleton is ε-PLL, another is IgG-XG-HRP whose skeleton is xanthan gum. We evaluated their performance by enzyme linked immunosorbent assay (ELISA) and immune histochemistry (IHC).Methods:(1) The synthetic route of IgG-PLL-HRP:First of all, we connected the hinge 2-(4-bromobutanamido)acetate with ε-PLL, purified through the silica gel column chromatography, and verificated by NMR and FTIR. And then, the ends of hinges were modified to form active ester. Finally, the antibodies and horseradish peroxidase (HRP) were conjugated to the ε-PLL skeleton at the same time, and the polymers were purified through sephacryl glucan gel chromatography separation. (2) The synthetic route of IgG-XG-HRP:First, we oxidated the adjacent hydroxyl of xanthan gum to form aldehyde with sodium periodate, and the product was verificated by BCA method. After that, the horseradish peroxidase was conjugated to the XG skeleton, and removed unbound HRP using sephacryl glucan gel chromatography. In the end, the antibodies were conjugated to XG-HRP, and the product was purified through sephacryl glucan gel chromatography separation.Results:(1) IgG-PLL-HRP was constructed successfully. But the performances of IgG-PLL-HRP in ELISA and IHC detection were similar to the commercially available IgG-HRP without polymer skeleton. (2) IgG-XG-HRP was constructed successfully. Compared to IgG-HRP, IgG-XG-HRP could improve the detective sensitivity with nearly 100 times. The performances of IgG-PLL-HRP in IHC was better than that of IgG-HRP.Conclusions:We successfully constructed the IgG-PLL-HRP and IgG-XG-HRP, and evaluated their performance through ELISA and IHC. The research provides some reference for future design of antibody and HRP polymer.