Production of polycolonal antibody against domain 2-4 of protective antigen of Bacillus anthracis in laboratory animals

Anthrax caused by bacillus anthracis، is an acute infectious disease that mostly occurs in herbivorous animals and in human. The vegetative format Bacillus anthracis is an exotoxin، which consists of three polypeptide: protective antigen (PA، 83 kDa)، lethal factor (LF، 90 kDa) and edema factor (EF، 89kDa). PA is considered as a primary immune gen for development of protective immunity against anthrax. The aim of this study was the production of antibody raised against domain 2-4 PA in lab animals. In this experimental study، domain 4-2 PA gene of plasmid pXOI with BamHI and HindIII restriction enzyme sites، amplified by PCR and were cloned and sub cloned in the vectors. Vector pET28a (+) was transformed to E. coli strain BL21 (DE3). After induction with IPTG، PA gene expression was observed. Purified protein was injected in mice and rabbits 4 times. Then the raised antibody was isolated from mice and rabbits sera and confirmed by ELISA. The pET28a (+) /domain 2-4 expression vector confirmed by end nuclease digestion، PCR and sequence analysis. The expression product domain 2-4 was confirmed by SDS-PAGE and Western blot. Antibody in mice and rabbits sera was confirmed by ELISA. Due to PA protein being immunogenic، it could be used for vaccine design and as a powerful mucosal adjuvant system.