Purification of 146s Antigen of Foot-and-Mouth Disease (FMD) Virus Serotypes Aby Using the Sucrose Gradient Procedure

Foot-and-Mouth disease (FMD) is an acute and contagious disease in domestic ruminants, which is currently the most economical viral disease that threatens the livestock industry. The virus that causes disease belongs to the Aphthovirus genus from the family of picornaviridae. This family contains seven serotypes and is about 30 nanometers in diameter and has no external membrane, similar to other picornaviruses.
In this study, we aimed to introduce the sucrose gradient procedure as a convenient method for purification of 146s antigens.
Sucrose gradient procedure (20% - 50%) was used for purification of 146s antigen of foot-and-mouth disease virus serotypes A. Then, preparation steps of the virus including concentration by polyethylene glycol, degreasing using trichloroethylene, centrifugation (30000 g for three hours) and washing the pellet using Tris (0.05 M) were performed. Spectrophotometer and nano-drop were used to measure the amount of the purified protein and purity evaluation, respectively. Moreover, dot blot assay was used for the confirmation of 146s antigen.
The pellet of the 146s antigen of FMD virus serotype A was developed at a concentration of sucrose 50%. Absorbance rate of the Foot-and-Mouth Disease virus serotypes A at wavelengths of 240, 259 and 280 nm was 1.238, 1.573 and 1.157, respectively. Moreover, the amount of 146s antigen at the same wavelengths was 163.416, 207.636 and 152.724 μg/mL, respectively. The amount of purified protein by nanodrop (ND-1000, the United States) was 0.275 mg/mL. The 146s antigen was observed with 26, 29 and 64 KDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and was confirmed by the dot blot assay.
The results exhibited that sucrose gradient procedure is a good method for purification of virus.