title: Identification and characterization of species-specific nanobodies for the detection of Listeria monocytogenes in milk.
abstract: Listeria monocytogenes (LM), one of the eight species belonging to the genus Listeria, is pathogenic for both humans and animals. In this study, two novel LM-specific clones, designated L5-78 and L5-79, were isolated from a phage display antibody library that was derived from the variable domain of heavy-chain antibodies (VHHs) of non-immunized alpaca. These two clones were expressed, purified, and characterized. Results showed that both isolated VHHs recognize three serotypes (1/2a, 1/2b, and 4b), which are responsible for more than 95% of documented human listeriosis cases. The recombinant VHHs possess high thermal stability, pH tolerance, and urea resistance. A sandwich enzyme-linked immunosorbent assay (ELISA) based on the VHH clone L5-79 and a monoclonal antibody was developed to detect LM in pasteurized milk, with a detection limit of 1 × 10(4) colony-forming units (CFU)/ml. These findings indicated that the species-specific VHHs could be directly isolated from the non-immunized library with a properly designed panning strategy and VHH could be a new source for possible diagnosis/detection of foodborne pathogens in food because it was shown to be highly specific and stable.
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China.
authors.forename: Zhui
authors.initials: Z
authors.lastname: Tu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China; Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China; Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047, China.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Li
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China; Jiangxi-OAI Joint Research Institute, 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; Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047, China. Electronic address: xuyang1951@163.com.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xu
authors.affiliation: College of Life Science, Jiangxi Normal University of Science and Technology, Nanchang 330013, China.
authors.forename: Na
authors.initials: N
authors.lastname: Hu
authors.affiliation: Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Tao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002161:Camelids, New World; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry; D008089:Listeria monocytogenes / Q000302:isolation & purification*; D008088:Listeriosis / Q000382:microbiology; D008892:Milk / Q000382:microbiology*; D008969:Molecular Sequence Data; D019151:Peptide Library; D016415:Sequence Alignment; D061905:Single-Domain Antibodies / Q000737:chemistry*; D013045:Species Specificity
chemical_list: D007143:Immunoglobulin Heavy Chains; D019151:Peptide Library; D061905:Single-Domain Antibodies
keywords: Enzyme-linked immunosorbent assay (ELISA); Listeria monocytogenes; Naive library; Phage display; Variable domain of heavy-chain antibodies (VHHs)