Xu Lingyuan, Abd El-Aty A M, Shim Jae-Han, Eun Jong-Bang, Lei Xingmei, Zhao Jing, Zhang Xiuyuan, Cui Xueyan, She Yongxin, Jin Fen, Zheng Lufei, Wang Jing, Jin Maojun, Hammock Bruce D
Institute of Quality Standard and Testing Technology for Agro-Products, Chinese Academy of Agricultural Sciences, Beijing 100081, China.
Department of Pharmacology, Faculty of Veterinary Medicine, Cairo University, Giza 12211, Egypt.
Foods. 2022 Jun 13;11(12):1726. doi: 10.3390/foods11121726.
This study provides the first design and synthetic protocol for preparing highly sensitive and specific atrazine (ATR) monoclonal antibodies (mAbs). In this work, a previously unreported hapten, 2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine, was designed and synthesized, which maximally exposed the characteristic amino group ATR to an animal immune system to induce the expected antibody. The molecular weight of the ATR hapten was 259.69 Da, and its purity was 97.8%. The properties of the anti-ATR mAb were systematically characterized. One 9F5 mAb, which can detect ATR, was obtained with an IC value (the concentration of analyte that produced 50% inhibition of ATR) of 1.678 µg/L for ATR. The molecular weight for the purified 9F5 mAb was approximately 52 kDa for the heavy chain and 15 kDa for the light chain. The anti-ATR mAb prepared in this study was the IgG type. The working range of the standard curve (IC (the concentration of analyte that produced 20% inhibition of ATR)IC (the concentration of analyte that produced 80% inhibition of ATR)) was 0.384 to 11.565 µg/L. The prepared anti-ATR mAb had high specificity, sensitivity, and affinity with low cross-reactivity. The prepared anti-ATR mAb could provide the core raw material for establishing an ATR immunoassay.
本研究提供了首个制备高灵敏度和特异性莠去津（ATR）单克隆抗体（mAb）的设计与合成方案。在本工作中，设计并合成了一种此前未报道的半抗原，即2-氯-4-乙氨基-6-异丙氨基-1,3,5-三嗪，它能使ATR的特征氨基最大限度地暴露于动物免疫系统，以诱导产生预期抗体。ATR半抗原的分子量为259.69 Da，纯度为97.8%。系统地表征了抗ATR单克隆抗体的特性。获得了一种能检测ATR的9F5单克隆抗体，其对ATR的IC值（产生50% ATR抑制作用的分析物浓度）为1.678 µg/L。纯化后的9F5单克隆抗体的重链分子量约为52 kDa，轻链分子量约为 15 kDa。本研究制备的抗ATR单克隆抗体为IgG型。标准曲线的工作范围（IC20（产生20% ATR抑制作用的分析物浓度）至IC80（产生80% ATR抑制作用的分析物浓度））为0.384至11.565 µg/L。所制备的抗ATR单克隆抗体具有高特异性、高灵敏度和高亲和力，交叉反应性低。所制备的抗ATR单克隆抗体可为建立ATR免疫分析提供核心原料。
