Biopolymers and Cell. 2012; 28(2): 141-148
BIOAFFINITY SORBENT BASED ON IMMOBILIZED PROTEIN A STAPHYLOCOCCUS AUREUS: DEVELOPMENT AND APPLICATION
Gorbatiuk O. B., 1Tsapenko M. V., 1, 2Pavlova M. V.1, 2Okunev O. V., 1, 2Kordium V. A.
Taras Shevchenko National University of Kyiv
64, Volodymyrska Str., Kyiv, Ukraine, 01033
1Institute of Molecular Biology and Genetics, NAS of Ukraine
150, Akademika Zabolotnogo Str., Kyiv, Ukraine, 03680
2State Institute of Genetic and Regenerative Medicine, NAMS of Ukraine
57/3, Chervonoarmiyska Str., Kyiv, Ukraine, 03150
Aim. The obtaining of bioaffinity sorbent based on the immobilized protein A of S. aureus (SPA) using two cellulose-binding domains (CBD), and its application for purification of antibodies. Methods. The DNA sequences encoding SPA and two CBD were genetically fused, expressed in the high-productive Escherichia coli system and the protein SPA-CBD2 was obtained in a soluble form. The SPA-CBD2 fusion protein was affinity immobilized on the microcrystalline cellulose. Results. Capacity of bioaffinity sorbent (1 mg SPA-CBD2/1 ml CC31-cellulose), dynamic capacity (3 mg mouse IgG/1 ml bioaffinity sorbent), efficiency and stability during prolonged storage were determined. The bioffinity sorbent was used for purification of antibodies. The purity of antibodies in eluted fractions was more than 95 %. The purified antibodies detected target antigens with a high sensitivity. Conclusions. The designed bioaffinity sorbent provides obtaining pure poly- and monoclonal antibodies in functionally active form and can be useful for the fractionation of mouse immunoglobulin G. |
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