Volume 39 - Issue 2 - 133 - 138

Affinity Separation and Characterization of IgG Subfragments by Fast Protein Liquid Chromatography with HiTrap_r Protein A Column

HiTrap_r Protein A Kolon İçeren Hızlı Protein Sıvı Kromatografisi ile IgG Altbirimlerinin Afinite Ayrılması ve Karakterizasyonu

It has been previously reported that the antibody molecules can be cleaved into fragments by the proteolytic enzymes and these fragments provide information to determine an antibody’s structure and function. Antibody fragments offer several advantages over intact antibodies for use in experimental applications and immunochemical techniques. These fragments have smaller sizes and higher chemical and physical resistances. In this study, papain was used for the digestion of human IgG and the resulting fragments were separated by Fast Protein Liquid Chromatography (FPLC). For this purpose, GE Healthcare HiTrap_r Protein A FF column was used, the resolution (Rs) and theoretical plate numbers (N) of the column were calculated. The unbound and bound fragments were collected by the Frac 920 fraction unit of the FPLC system and the collected fragments were analysed by enzyme-linked immunosorbent assay (ELISA) and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). 

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