2. Academic Validation
  3. Binding affinity-guided design of a highly sensitive noncompetitive immunoassay for small molecule detection

Binding affinity-guided design of a highly sensitive noncompetitive immunoassay for small molecule detection

  • Food Chem. 2021 Jul 30:351:129270. doi: 10.1016/j.foodchem.2021.129270.
Yuchen Bai 1,  Yahui Wang 2,  Qiang Li 1,  Leina Dou 1,  Minggang Liu 1,  Shibei Shao 1,  Jianyu Zhu 1,  Jianzhong Shen 1,  Zhanhui Wang 1,  Kai Wen 3,  Wenbo Yu 4
Affiliations

Affiliations

  • 1 College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal Derived Food Safety, Beijing Laboratory for Food Quality and Safety, 100193 Beijing, People's Republic of China.
  • 2 College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal Derived Food Safety, Beijing Laboratory for Food Quality and Safety, 100193 Beijing, People's Republic of China; Agricultural Information Institute, Chinese Academy of Agricultural Sciences, 100081 Beijing, People's Republic of China.
  • 3 College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal Derived Food Safety, Beijing Laboratory for Food Quality and Safety, 100193 Beijing, People's Republic of China. Electronic address: wenkai@cau.edu.cn.
  • 4 College of Veterinary Medicine, China Agricultural University, Beijing Key Laboratory of Detection Technology for Animal Derived Food Safety, Beijing Laboratory for Food Quality and Safety, 100193 Beijing, People's Republic of China. Electronic address: yuwenbo@cau.edu.cn.
PMID: 33640770 DOI: 10.1016/j.foodchem.2021.129270
Abstract

Small molecules are immunochemically classified as Hapten that lacking of at least two epitopes, usually using competitive format for establishing immunoassays. However, theoretically, noncompetitive immunoassay format is more sensitive and has a wider analytical range. In the present study, a novel Hapten of halofuginone was synthesized and used to produce a monoclonal antibody (mAb). By analyzing the binding kinetics, we found that the affinity of analyte-enzyme to mAb was much greater than that of analyte, which could result in a low sensitivity of competitive assay format. Based on this, we established a novel noncompetitive immunoassay by using a replacement approach. The noncompetitive format has obvious advantages in sensitivity and analytical range, which promoted approximately 3.5- and 5-fold, respectively, compared to the competitive immunoassay. Ultimately, the newly designed noncompetitive immunoassay in this work will provide insights as well as alternative method to traditional small molecule competitive assays.

Keywords

Affinity; Halofuginone; Noncompetitive immunoassay; Small molecules.

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