title: Sensitive detection of Escherichia coli O157:H7 based on cascade signal amplification in ELISA.
abstract: In this study, cascade signal amplification in ELISA involving double-antibody sandwich ELISA and indirectly competitive ELISA was established to sensitively detect Escherichia coli O157:H7. In the double-antibody sandwich ELISA, a complex was formed comprising anti-E. coli O157:H7 polyclonal antibody, E. coli O157:H7, biotinylated anti-E. coli O157:H7 monoclonal antibody, streptavidin, and biotinylated β-lactamase. Penicillin solution was then added into the ELISA well and hydrolyzed by β-lactamase. Afterward, the penicillin solution was transferred to indirectly competitive ELISA. The concentration of penicillin can be sensitively detected in indirectly competitive ELISA. In the cascade signal amplification system, increasing the amount of added E. coli O157:H7 resulted in more β-lactamase and less penicillin. The detection sensitivity of E. coli O157:H7, which was 20cfu/mL with the cascade signal amplification in ELISA, was 1,000-fold higher than that of traditional ELISA. Furthermore, the novel method can be used to detect E. coli O157:H7 in milk (2cfu/g). Therefore, this new signaling strategy will facilitate analyses of highly sensitive foodborne pathogens.
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Shan
authors.affiliation: Jiangxi Province Centers for Disease Control and Prevention, Nanchang 330096, China.
authors.forename: Daofeng
authors.initials: D
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Guo
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Songsong
authors.initials: S
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Kai
authors.initials: K
authors.lastname: Luo
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Liming
authors.initials: L
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Yonghua
authors.initials: Y
authors.lastname: Xiong
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China. Electronic address: talktolaiwh@163.com.
authors.forename: Weihua
authors.initials: W
authors.lastname: Lai
mesh_terms: D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D004927:Escherichia coli Infections; D019453:Escherichia coli O157 / Q000276:immunology*; D008892:Milk; D001618:beta-Lactamases
chemical_list: D001618:beta-Lactamases
keywords: ELISA; Escherichia coli O157:H7; milk; signal amplification