Purification of the Specific Antibody of Foot-and-Mouth Disease Virus Serotype A by Ion Exchange Chromatography

Foot-and-mouth disease (FMD) is an acute and contagious disease in domestic ruminants, which is currently the most economical viral disease that threatens livestock industry. The virus that causes the disease is belongs to Aphthovirus genus from the picornaviridae family. This family contains seven serotypes and is about 30 nanometers in diameter and no external membrane similar to other picornaviruses.
The current study aimed to introduce ion exchange chromatography as a convenient method to purify specific antibody against 146S antigen.
The study was performed in vaccine and serum research of Razi institute, Tehran, Iran, after purification of 146S antigen of foot-and-mouth disease virus serotypes A using sucrose gradient procedure; two Guinea pigs were immunized with 30 µg 146S antigen combined with complete Freund’s adjuvant (CFA) and booster with incomplete Freund’s adjuvant (IFA) according to the protocol. After bleeding and serum obtaining, agglutination assay, dot blot and the enzyme linked immunosorbent assay (ELISA) were used to confirm specific antibody against 146S. Ion exchange chromatography was used to purify specific antibody against 146S antigen. Finally, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) electrophoresis was used to evaluate the purification.
The interaction of 146S antigen with the immunized rabbit resulted in agglutination reaction. Assaying with the heterologous antigen showed negative result that confirmed the production of specific antibody in the rabbit. In dot blot, presence of brown spots was confirmed by binding specific antibody with 146S antigen. The result of ELISA showed that each antigen of different serotypes reacted better with homolog antibody. Since IgG containing positive charge and 25, 2-diethylaminoethyl (DEAE) Cephadex gel in ion exchange chromatography also had positive charge, IgG was eluted at the first step by buffer (pH = 9) and the other proteins were eluted by buffer at different pH levels. The absorbance amount of IgG was 0.197 that showed IgG = 0.725 mg/mL.
With respect to the results as well as the speed of ion exchange chromatography, this method is advised to purify antibodies.