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Published on: November 17, 2023
Phage display-derived single-chain variable fragment antibody for highly sensitive and specific detection of
Jiadi Sun1, Yan Lai2, Wancheng Ma2
1School of Food Science and Technology, International Joint Laboratory on Food Safety, Synergetic Innovation Center of Food Safety and Quality Control, Jiangnan University, Wuxi, Jiangsu, 214122, PR China; Key Laboratory of Screening, Prevention, and Control of Food Safety Risks, State Administration for Market Regulation, Wuxi, Jiangsu, 214122, PR China; Institute of Future Food Technology, JITRI, Yixing, Jiangsu, 214200, PR China; Key Laboratory of Biotechnology and Bioresources Utilization of Ministry of Education, School of Life Sciences, Dalian Minzu University, Dalian, 116600, PR China.
Abstract:
Thiamethoxam (TMX), as a widely used pesticide, poses potential risks to food safety due to residual contamination, underscoring the need for highly sensitive recognition elements for its detection. This study constructed a phage-displayed immune antibody library via mouse immunization with a titer of 1.6 × 1010 PFU/mL. Following four rounds of biopanning, a soluble single-chain variable fragment (scFv) antibody was isolated, which can specifically recognize TMX. Based on this antibody, an indirect competitive ELISA (ic-ELISA) and colloidal gold immunochromatographic assay (CG-ICA) were established. Under the optimal conditions for ic-ELISA, the IC50 value was 25.95 ng/mL, with a linear response range of 8.13 ng/mL - 137.97 ng/mL. Additionally, the scFv showed acceptable specificity against other structurally related neonicotinoids. In spiked real samples, average recoveries ranged from 71.14% to 110.49% with coefficients of variation of 2.22%-7.92%, demonstrating good accuracy and precision. Furthermore, the visual limit of detection (vLOD) for the optimized CG-ICA was 0.06 mg/kg - 0.10 mg/kg. This work offers theoretical insight and technical assistance for developing scFv antibodies against TMX residues in various samples.
