Shen Hong, Li Chao, Sun Han, Chen Wanqin, Chen Bilian, Yi Yu, Mei Jianfeng, Zhang Yanlu, Ying Guoqing
College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, China.
Department of Biological Inspection, Zhejiang Institute for Food and Drug Control, Hangzhou, China.
Front Nutr. 2022 May 16;9:910876. doi: 10.3389/fnut.2022.910876. eCollection 2022.
An anti-diclazuril monoclonal antibody (mAb) was developed for use in enzyme-linked immunosorbent assay (ELISA)-based detection of diclazuril with high sensitivity and specificity, which can be used to measure anti-coccidial drug residues. The anti-diclazuril mAb had a half-maximal inhibitory concentration of 0.449-0.517 ng/mL. The mAb cross-reactivity with toltrazuril, toltrazuril 18 sulfone, clozaril, monesin, madurmycin, and salinomycin was very minimal (< 0.1%). The detection limit of the ELISA using this mAb was 0.10 ng/mL and the sensitivity was 0.05 ng/mL. A standard curve generated in the range of 0.05-16.2 ng/mL had a linear correlation coefficient value of ≥ 0.99. The average recoveries of diclazuril from chicken and duck samples ranged from 85.0 to 102.5%.Intra- and inter-assay coefficients of variation ranged from 5.9 to 8.5% and 9.2 to 12.6%, respectively. Using the International Immunogenetics Information System, the VH domain of the mAb was found to be encoded by an IGHV3 family gene and had the following complementarity determining region (CDR) sequences: GFTFSRY (CDR1), SRGGS (CDR2), and GDDNYAFAY (CDR3). The VL domain was encoded by an IGKV1 family gene and had the following CDR sequences: KSSQSLLNSRTRKNYLA (CDR1), WASTRES (CDR2), and KQSYNLHT (CDR3). This study provides a method to generate anti-diclazuril mAbs and determine their variable region sequences. The diagnostic ELISA developed using this mAb may drive additional studies on the monitoring and detection of food and veterinary drug residues.
开发了一种抗地克珠利单克隆抗体（mAb），用于基于酶联免疫吸附测定（ELISA）的高灵敏度和特异性地克珠利检测，可用于测量抗球虫药物残留。抗地克珠利单克隆抗体的半数抑制浓度为0.449 - 0.517 ng/mL。该单克隆抗体与托曲珠利、托曲珠利18 - 砜、氯氮平、莫能菌素、马杜霉素和盐霉素的交叉反应性非常低（< 0.1%）。使用该单克隆抗体的ELISA检测限为0.10 ng/mL，灵敏度为0.05 ng/mL。在0.05 - 16.2 ng/mL范围内生成的标准曲线的线性相关系数值≥0.99。地克珠利在鸡和鸭样品中的平均回收率为85.0%至102.5%。批内和批间变异系数分别为5.9%至8.5%和9.2%至12.6%。使用国际免疫遗传学信息系统，发现该单克隆抗体的重链可变区（VH）由IGHV3家族基因编码，并具有以下互补决定区（CDR）序列：GFTFSRY（CDR1）、SRGGS（CDR2）和GDDNYAFAY（CDR3）。轻链可变区（VL）由IGKV1家族基因编码，并具有以下CDR序列：KSSQSLLNSRTRKNYLA（CDR1）、WASTRES（CDR2）和KQSYNLHT（CDR3）。本研究提供了一种产生抗地克珠利单克隆抗体并确定其可变区序列的方法。使用该单克隆抗体开发的诊断ELISA可能推动食品和兽药残留监测与检测的更多研究。
