猪肉中2-(三氟甲基)吩噻嗪的Ic-ELISA检测  

Detection of 2 -( trifluoromethyl ) phenothiazine in pork by Ic- ELISA

作　　者：刘广兴 韩晓利 李昳晴 王庭欣[1] LIU Guangxing;HAN Xiaoli;LI Yiqing;WANG Tingxin(Hebei Key Laboratory of Energy Metering and Safety Testing Technology,National&Local Joint Engineering Research Center of Metrology Instrument and System,College of Quality and Technical Supervision,Hebei University,Baoding 071002,China)

机构地区：[1]河北大学质量技术监督学院,计量仪器与系统国家地方联合工程研究中心,河北省能源计量与安全检测技术重点实验室(筹),河北保定071002

出　　处：《河北大学学报（自然科学版）》2023年第6期638-645,共8页Journal of Hebei University(Natural Science Edition)

基　　金：河北省重点研发计划项目(18275502D)。

摘　　要：为了快速检测猪肉中的镇静剂2-(三氟甲基)吩噻嗪(2-(trifluoromethyl)phenothiazine,TFPTZ),分别用偶氮苯甲酸法、混合酸酐法合成了半抗原、完全抗原,通过免疫新西兰大耳白兔获得了多克隆抗体,采用间接竞争酶联免疫方法(Ic-ELISA)对猪肉中TFPTZ进行了定量分析,实验条件:包被抗原浓度1.25μg/mL,多克隆抗体稀释16000倍,37℃包被3 h,封闭液为质量分数1%的牛血清白蛋白(BSA),TFPTZ稀释液为含体积分数10%的DMSO的PBS,竞争反应60 min.该方法相关系数R^(2)为0.9991,IC_(50)为1.5002μg/L,最低检测限IC_(10)为0.2841μg/L,线性(IC_(20)~IC_(80))为0.4307~5.2252μg/L,与5种TFPTZ结构类似物的交叉反应率小于1.2%,样品添加回收率为93.26%~109.13%,变异系数小于6%,实际样品检测结果与高效液相色谱法测定结果高度一致,表明建立的快速检测猪肉中TFPTZ的Ic-ELISA方法具有良好的准确度,适用于快速检测猪肉中TFPTZ的残留量.In order to quickly analyze the sedative 2-(trifluoromethyl)phenothiazine(TFPTZ)in pork,haptens and complete antigens were synthesized using the azobenzoic acid method and the mixed anhydride method,respectively.Polyclonal antibodies were obtained by immunizing New Zealand white rabbits.Indirect competitive enzyme linked immunosorbent assay(Ic-ELISA)was used to quantitatively analyze TFPTZ in pork.The experimental conditions were as follows:the concentration of the coated antigen was 1.25μg/mL,and the polyclonal antibody was diluted 16000 times,Coated at 37℃for 3 h,the blocking solution was 1%BSA of mass fraction,and the TFPTZ diluent is PBS of DMSO with a volume fraction of 10%.The competitive reaction lasted 60 minutes.The correlation coefficient R^(2) of this method was 0.9991,and the IC_(50) was 1.5002μg/L,minimum detection limit IC_(10) is 0.2841μg/L,linear range(IC_(20)~IC_(80))0.4307~5.2252μg/L,the cross reaction rate with five TFPTZ structural analogues is less than 1.2%,the sample addition recovery rate is 93.26%~109.13%,addition recovery rate is 93.26%~109.13%,and the coefficient of variation is less than 6%.The actual sample detection results are highly consistent with the results determined by high-performance liquid chromatography,indicating that the established Ic-ELISA detection method of TFPTZ has good accuracy and is suitable for rapid detection of TFPTZ residues in pork.

关 键 词：2-(三氟甲基)吩噻嗪 抗原合成 间接竞争酶联免疫方法(Ic-ELISA) 

分 类 号：Q5-33[生物学—生物化学]