摘要
建立一种3-氨基-2-恶唑烷酮(3-amino-2-oxazolidinone,AOZ)夹心免疫检测方法。通过1,6-己二醇连接2-硝基-4-羧基苯甲醛和生物素合成针对AOZ的新型衍生试剂。在样品前处理时加入衍生试剂可对AOZ进行衍生,衍生产率为89%。在酶标板上包被抗AOZ单克隆抗体并以辣根过氧化物酶标记的亲和素或抗生物素抗体作为第二结合物可实现AOZ的夹心酶联免疫吸附检测(enzyme-linked immunosorbnent assay,ELISA)。从实际应用的角度,得到双抗夹心和抗体-亲和素夹心模式下两个表位的极限距离,分别为12Å和13Å,理想距离分别为16Å和17Å。在双抗夹心和抗体-亲和素夹心模式下的检出限分别达到1.8 pg/mL和0.8 pg/mL(以AOZ质量浓度计),相对于竞争ELISA,其灵敏度最高提高了25倍。平均回收率为73%~85%,平均相对标准偏差为9.0%。
In the present study,a sandwich enzyme-linked immunosorbent assay(ELISA)for 3-amino-2-oxazolidnone(AOZ)was developed.A novel derivatizing agent for AOZ was synthesized by linking 2-nitro-4-carboxybenzaldehyde and biotin through 1,6-hexanediol.During the sample pretreatment,the agent was added for AOZ derivatization with an efficiency of 89%.The monoclonal antibody(McAb)against AOZ was coated on the microplate and horseradish peroxidase-labeled avidin or anti-biotin antibody was used as a secondary conjugate in the ELISA method.From a practical perspective,the limit distances between the two epitopes in the double-antibody sandwich and antibody-avidin sandwich modes were 12 and 13Å,respectively,and the ideal distances were 16 and 17Å,respectively.The detection limits of the double-antibody sandwich and antibody-avidin sandwich modes were 1.8 and 0.8 pg/mL for AOZ,respectively.Compared with competitive ELISA,a 25-fold improvement in the sensitivity of the developed ELISA method was achieved.The average recoveries and average relative standard deviations(RSDs)were 73%–85%and 9.0%,respectively.
作者
张世伟
吴会玲
周迎春
王炳志
杨星星
杜业刚
汤璐
冯荣虎
郭继平
ZHANG Shiwei;WU Huiling;ZHOU Yingchun;WANG Bingzhi;YANG Xingxing;DU Yegang;TANG Lu;FENG Ronghu;GUO Jiping(Shenzhen Academy of Metrology&Quality Inspection,Shenzhen 518000,China;Shenzhen Bioeasy Biotechnology Co.Ltd.,Shenzhen 518000,China)
出处
《食品科学》
EI
CAS
CSCD
北大核心
2024年第6期271-276,共6页
Food Science
基金
深圳市科技计划项目(KCXFZ20230731092859013
JCYJ20210324141804012
JCYJ20210324141801003)。
