Development of ELISA based on Bacillus anthracis capsule biosynthesis protein CapA for naturally acquired antibodies against anthrax
Fig 2
Expression and purification of CapA322.
(A) Genetic map of Bacillus anthracis capsule-coding operon revealing the CapA322, C-terminus region (amino-acid 322–411) of CapA used in antigen preparation. (B) Coomassie brilliant blue staining (CBB) and (C, D, and G) Western blotting of the fractions collected after affinity batch purification. In Western blotting, proteins were probed with different horse sera: (C) horse hyperimmunized anti-B. anthracis serum (PC1); (D) naive horse serum (NC1); (G) vaccinated horse serum (Vac_H3D21). Lane 1, Mw, molecular weight marker (in kDa); lane 2, the supernatant fraction applied to affinity beads (GST-CapA322: 37 kDa); lane 3, beads bound; lane 4, elute after treatment with PreScission Protease (CapA322: 11 kDa). (E and F) CBB and Western blotting of the fractions collected during the cation exchange process. Lane 1, Mw, molecular weight marker (in kDa); lane 2, sample loaded onto the cation exchange chromatography column; lanes 3 and 4 flow-throughs; lane 5, the eluted protein. Host cell-derived contaminants indicated as Escherichia coli’s protein.