title: Anti-idiotypic nanobody: A strategy for development of sensitive and green immunoassay for Fumonisin B₁.
abstract: Nanobodies that are small and thermally stable, as well as have high expression level, are leading alternative to produce anti-idiotypic antibodies. These antibodies have the advantage of replacing mycotoxins and their conjugates for immunoassays. In this work, anti-fumonisin B1 (FB1) monoclonal antibody (mAb) was used as the target for biopanning from a naïve alpaca nanobody (Nb) phage display library. After three cycles of panning, one anti-idiotypic nanobody (Ab2β Nb) was isolated and subjected to a Nb-ELISA for the detection of FB1. Surface plasmon resonance was used to analyze the reaction kinetics between the Ab2β Nb and anti-FB1 mAb. The developed assay showed a half inhibitory concentration (IC50) of 0.95±0.12 ng/mL, a limit of detection of 0.15 ng/mL, a linear range of 0.27-5.92 ng/mL, and a low cross-reactivity toward FB2 of 4.93%. The sensitivity was enhanced approximately 20-fold compared with that of the chemosynthetic FB1-BSA conjugates. The equilibrium dissociation constant (KD) measured for Ab2β Nb: anti-FB1 mAb was 164.6 nM. The assay was compared with conventional ELISA (the commercial ELISA kit), and the results indicated the reliability of Ab2β Nb replacing the antigen-carrier protein conjugates. The use of biotechnology in developing the surrogate is an ideal strategy for replacing conventional synthesized antigens.
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Mei
authors.initials: M
authors.lastname: Shu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China. Electronic address: xuyang@ncu.edu.cn.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Key Lab for Agricultural Products Processing and Quality Control of Nanchang City, Jiangxi Agricultural University, Nanchang 330045, Jiangxi, People's Republic of China.
authors.forename: Dan
authors.initials: D
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Xing
authors.initials: X
authors.lastname: Liu
authors.affiliation: Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Li
authors.affiliation: Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Qinghua
authors.initials: Q
authors.lastname: He
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Zhui
authors.initials: Z
authors.lastname: Tu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Yulou
authors.initials: Y
authors.lastname: Qiu
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Yanwei
authors.initials: Y
authors.lastname: Ji
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China; Sino-Germany Joint Research Institute, Nanchang University, No. 235 Nanjing East Road, Nanchang 330047, People's Republic of China.
authors.forename: Xianxian
authors.initials: X
authors.lastname: Wang
mesh_terms: D000888:Antibodies, Anti-Idiotypic / Q000276:immunology*; D037341:Fumonisins / Q000032:analysis* / Q000276:immunology; D007118:Immunoassay; D061905:Single-Domain Antibodies / Q000276:immunology*
chemical_list: D000888:Antibodies, Anti-Idiotypic; D037341:Fumonisins; D061905:Single-Domain Antibodies; C056933:fumonisin B1
keywords: Anti-idiotypic antibodies; Nanobodies; Phage display library