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ABSTRACT
Immunosorbents in immunoaffinity chromatography (IAC) are prepared by immobilizing expensive antibodies without guidelines for ensuring the best coupling efficiencies, and avoiding low binding capacities. Covalent immobilization of antibodies on N-hydroxysuccinimide (NHS)-activated Sepharose 4 Fast Flow resin was optimized using human IgG via full factorial design with incubation times (4, 9, 14, 19 and 24 h), temperatures (4°C and 20°C) and coupling reaction buffers (sodium bicarbonate and triethanolamine). The best coupling efficiency (CE) (83.4 ± 8.7%) was reached with triethanolamine buffer, 14 h and 4°C. Comparison of antibody isotypes (IgG or IgM) by a nested factorial analysis suggested that antibodies in the IgG isotype presents the best coupling efficiency.
Declaration of interest
The authors have declared no conflicts of interest.
Funding
The authors acknowledge the financial support of Tecnológico de Monterrey through the Bioprocess and Synthetic Biology Strategic Focus Group (0821C01004) and CONACyT for the funding of Project 242286. Additionally, Luis Alberto Mejía-Manzano thanks CONACyT for grant number 252731.