title: An affinity improved single-chain antibody from phage display of a library derived from monoclonal antibodies detects fumonisins by immunoassay.
abstract: Fumonisin B analogs, particularly FB1, FB2, and FB3, are major mycotoxins found in cereals. Single-chain fragment variable (scFv) antibodies represent a promising alternative immunoassay system. A phage-displayed antibody library derived from four monoclonal antibodies (mAbs) generated against FB1 was used to screen high binding affinity scFv antibodies; the best candidate was designated H2. Surface plasmon resonance measurements confirmed that the H2 scFv displayed a 82-fold higher binding affinity than its parent mAb. Direct competitive enzyme-linked immunosorbent assay demonstrated that the H2 antibody could competitively bind to free FB1, FB2, and FB3, with an IC50 of 0.11, 0.04, and 0.10 μM, respectively; it had no cross-reactivity to deoxynivalenol, nivalenol and aflatoxin. Validation assays with naturally contaminated samples revealed a linear relationship between the H2 antibody-based assay results and chemical analysis results, that could be expressed as y=1.7072x+5.5606 (R(2)=0.8883). Homology modeling of H2 revealed a favorable binding structure highly complementary to the three fumonisins. Molecular docking analyses suggested that the preferential binding of the H2 scFv to FB2 was due to the presence of a hydrogen radical in its R1 position, leading to a proper electrostatic matching and hydrophobic interaction. The H2 scFv antibody can be used for the rapid, accurate, and specific detection of fumonisin contamination in agricultural samples.
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China; College of Biology and Engineering, Guizhou Medical University, Guiyang 550004, China.
authors: Zu-Quan
authors: ZQ
authors: Hu
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: He-Ping
authors: HP
authors: Li
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: Ping
authors: P
authors: Wu
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: Ya-Bo
authors: YB
authors: Li
authors: College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: Zhu-Qing
authors: ZQ
authors: Zhou
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: Jing-Bo
authors: JB
authors: Zhang
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
authors: Jin-Long
authors: JL
authors: Liu
authors: Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070, China; College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China; National Center of Plant Gene Research (Wuhan), Wuhan 430070, China. Electronic address: yucailiao@mail.hzau.edu.cn.
authors: Yu-Cai
authors: YC
authors: Liao
mesh_terms: D000915:Antibody Affinity; D002460:Cell Line; D003429:Cross Reactions; D037341:Fumonisins / Q000032:analysis* / Q000276:immunology; D006825:Hybridomas / Q000166:cytology; D007118:Immunoassay; D007700:Kinetics; D062105:Molecular Docking Simulation; D019151:Peptide Library; D011487:Protein Conformation; D017421:Sequence Analysis; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology*; D012995:Solubility
chemical_list: D037341:Fumonisins; D019151:Peptide Library; D057127:Single-Chain Antibodies
keywords: Binding kinetics; Direct competitive enzyme-linked immunosorbent assay; Fumonisin; Monoclonal recombinant antibody; Phage display