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Identification and characterization of species-specific nanobodies for the detection of Listeria monocytogenes in milk

Author:
Tu, Zhui, Chen, Qi, Li, Yanping, Xiong, Yonghua, Xu, Yang, Hu, Na, Tao, Yong
Source:
Analytical biochemistry 2016 v.493 pp. 1-7
ISSN:
0003-2697
Subject:
Listeria monocytogenes, alpacas, bacteriophages, clones, detection limit, enzyme-linked immunosorbent assay, food pathogens, humans, listeriosis, milk, monoclonal antibodies, pH, pasteurized milk, serotypes, thermal stability, urea
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 × 104 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.
Agid:
5285698