title: Quantitative Determination of Lethal Toxin Proteins in Culture Supernatant of Human Live Anthrax Vaccine Bacillus anthracis A16R.
abstract: Bacillus anthracis (B. anthracis) is the etiological agent of anthrax affecting both humans and animals. Anthrax toxin (AT) plays a major role in pathogenesis. It includes lethal toxin (LT) and edema toxin (ET), which are formed by the combination of protective antigen (PA) and lethal factor (LF) or edema factor (EF), respectively. The currently used human anthrax vaccine in China utilizes live-attenuated B. anthracis spores (A16R; pXO1+, pXO2-) that produce anthrax toxin but cannot produce the capsule. Anthrax toxins, especially LT, have key effects on both the immunogenicity and toxicity of human anthrax vaccines. Thus, determining quantities and biological activities of LT proteins expressed by the A16R strain is meaningful. Here, we explored LT expression patterns of the A16R strain in culture conditions using another vaccine strain Sterne as a control. We developed a sandwich ELISA and cytotoxicity-based method for quantitative detection of PA and LF. Expression and degradation of LT proteins were observed in culture supernatants over time. Additionally, LT proteins expressed by the A16R and Sterne strains were found to be monomeric and showed cytotoxic activity, which may be the main reason for side effects of live anthrax vaccines. Our work facilitates the characterization of anthrax vaccines components and establishment of a quality control standard for vaccine production which may ultimately help to ensure the efficacy and safety of the human anthrax vaccine A16R.
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. zaixiaodong@163.com.
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Zai
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. justforhere@126.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. 18701522421@163.com.
authors.forename: Ju
authors.initials: J
authors.lastname: Liu
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. liujieer2015@163.com.
authors.forename: Jie
authors.initials: J
authors.lastname: Liu
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. byebyelll@163.com.
authors.forename: Liangliang
authors.initials: L
authors.lastname: Li
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. yinying1028@sina.cn.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yin
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. fuling3436@163.com.
authors.forename: Ling
authors.initials: L
authors.lastname: Fu
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. xujunjie@sina.com.
authors.forename: Junjie
authors.initials: J
authors.lastname: Xu
authors.affiliation: Laboratory of Vaccine and Antibody Engineering, Beijing Institute of Biotechnology, Beijing 100071, China. cw0226@foxmail.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
mesh_terms: D000818:Animals; D022122:Anthrax Vaccines; D000942:Antigens, Bacterial / Q000378:metabolism* / Q000633:toxicity; D001408:Bacillus anthracis / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism* / Q000633:toxicity; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D051379:Mice; D014613:Vaccines, Attenuated
chemical_list: D022122:Anthrax Vaccines; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D014613:Vaccines, Attenuated; C030325:anthrax toxin
keywords: A16R; Bacillus anthracis; anthrax toxin; anthrax vaccine; lethal factor; lethal toxin; protective antigen; quantitative determination