花生致敏蛋白Ara h 1双抗体夹心ELISA法的建立  

Establishment of double antibody sandwich ELISA for peanut allergenic protein Ara h 1

作　　者：王耀[1] 陈曦[1] 武汉良 卫星 张国庆 张淑霞 刘胜男 WANG Yao;CHEN Xi;WU Han-liang;WEI Xing;ZHANG Guo-qing;ZHANG Shu-xia;LIU Sheng-nan(College of Food and Bioengineering,Henan University of Science&Technology,Luoyang,Henan 471023,China;Clinical Laboratory,No.989 Military Hospital,Joint Logistics Support Force of PLA,Luoyang,Henan 471031,China;Sanmenxia Inspection and Quarantine Center,Sanmenxia,Henan 472000,China;Technology Center of Zhengzhou Customs,Zhengzhou,Henan 450002,China;Sanmenxia Customs District,Sanmenxia,Henan 472000,China)

机构地区：[1]河南科技大学食品与生物工程学院,河南洛阳471023 [2]中国人民解放军联勤保障部队第九八九医院检验科,河南洛阳471031 [3]三门峡市检验检疫中心,河南三门峡472000 [4]郑州海关技术中心,河南郑州450002 [5]三门峡海关,河南三门峡472000

出　　处：《食品与机械》2020年第7期59-62,113,共5页Food and Machinery

基　　金：海关总署科技计划项目(编号:2017IK284,2017IK283);河南省高等学校重点科研项目(编号:18B550001)。

摘　　要：为建立用于花生致敏蛋白Ara h 1快速、灵敏、特异检测的双抗体夹心ELISA法,以Ara h 1鼠源单抗(mAb)为捕获抗体,以Ara h 1兔源多抗(pAb)为检测抗体,通过棋盘法优化抗体工作浓度建立方法,并对方法的灵敏度、特异性、准确度、稳定性等检测特性进行鉴定。结果表明,双抗体夹心ELISA法的mAb和pAb最佳工作浓度分别为1∶10000稀释和1∶8000稀释;ELISA标准曲线线性范围为4~256 ng/mL,检测限为4.16 ng/mL;样品添加回收试验的平均回收率为85.9%~94.5%,且该方法与其他常见致敏蛋白无交叉反应;并能在4℃条件下避光密封保存90 d内保持检测结果稳定。说明所建立的双抗体夹心ELISA法灵敏特异、准确稳定,能够用于花生致敏蛋白Ara h 1的快速筛查。In order to establish a double antibody sandwich ELISA method for the rapid,sensitive and specific detection of peanut allergenic protein Ara h 1.Taking Ara h 1 murine monoclonal antibody(mAb)as capture antibody,and Ara h 1 rabbit polyclonal antibody(pAb)as detection antibody,the checkerboard method was used to optimize the antibody working concentration to establish the method,and the detection characteristics were identified,including sensitivity,specificity,accuracy and stability of the method.Results:The optimal working concentrations of mAb and pAb were 1∶10000 and 1∶8000 dilution,respectively.The linear range of the ELISA standard curve was 4~256 ng/mL,and the limit of detection was 4.16 ng/mL.The average recovery ranged from 85.9%to 94.5%,and there was no cross reaction with other common allergenic proteins.In addition,the detection results stable within 90 days under the condition of 4℃in the dark and sealed.The established double antibody sandwich ELISA method is sensitive,specific,accurate and stable,which can be used for the rapid screening of peanut allergenic protein Ara h 1.

关 键 词：花生 Ara h 1蛋白 双抗体夹心法 酶联免疫吸附 

分 类 号：TS201.21[轻工技术与工程—食品科学]