Design of bio-inspired silica-encapsulated protein A for improved immunoprecipitation assays

초록

Staphylococcus aureus Protein A (SpA) has a high affinity to the Fc region of antibodies (Abs), and SpA-immobilized matrices are widely used for Ab purification or immunoprecipitation (IP) assays. Here, we employed a bio-inspired silica-encapsulation method to improve the Ab-binding ability of an SpA-immobilized matrix. Two silica-forming peptides (SFPs), namely R5 and EctP1, were separately introduced at the C-terminus of SpA to generate two recombinant fusion proteins (SpA-SFPs) with auto-silicifying abilities. When SpA-SFPs were employed as Ab-binders on a solid surface (96-well plate), they showed an effective Ab-binding ability and a better performance than intact SpA. A high binding ability was observed even when an SFP-mediated SpA silica matrix (SpA-SFP@SiO2) was prepared. SpA-SFP@SiO2 showed a higher performance than commercially obtained SpA-Agarose particles and no loss of matrices. Moreover, in IP assays, SpA-SFP@SiO2 showed an approximately 300% higher precipitation of target protein than the commercial SpA-Agarose product when a small amount of cell lysate was used. These findings demonstrated that SpA-SFP could be useful for the development of an efficient immunoassay system. (C) 2017 Elsevier B.V. All rights reserved.

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