基于衍生化3-氨基-2-恶唑烷酮的夹心酶联免疫分析法研究

摘要/Abstract

摘要： 呋喃唑酮是一种经常被滥用的硝基呋喃类抗生素。其代谢物3-氨基-2-恶唑烷酮(AOZ）分子量仅为102Da。本研究建立一种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了针对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，衍生化率92%。在酶标板上包被抗AOZ单克隆抗体并使用辣根过氧化物酶（HRP）标记的亲和素或抗生物素抗体作为第二结合物可实现AOZ的夹心酶联免疫检测（ELISA）。从实际应用的角度，得到了双抗夹心和抗体-亲和素夹心模式下两个表位的极限距离分别为12 ?和13 ?，理想距离均为≥17 ?。在双抗夹心和抗体-亲和素夹心模式下的最低检出限分别达到1.8 pg/mL和0.8 pg/mL（以AOZ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%。

关键词: 呋喃唑酮, 小分子, 夹心免疫分析

Abstract: urazolidone [N-(5-nitro-2-furfurylidenenamino)-2-oxazolidinone] is a member of the nitrofuran antibiotics family which are often abused in animal farming. Its metabolite, 3-Amino-2-oxazolidinone (AOZ), is a small molecule compound with a molecular weight of 102 Da. In the present study, a sandwich enzyme-linked immunosorbent assay (ELISA) for AOZ was developed. A novel derivatizing agent for AOZ was synthesized by linking 2-nitro-4-carboxybenzaldehyde and biotin with 1,6-hexanediol; the agent was added during sample pretreatment for AOZ derivatization with 92% yield. Sandwich ELISA for AOZ was developed by coating the ELISA plate with anti-AOZ monoclonal antibody and using horseradish peroxidase-labeled avidin or anti-biotin antibody as a secondary conjugate. From the perspective of application, the limit distances of the two epitopes under the double-antibody sandwich and antibody-avidin sandwich modes were 12 and 13 ?, respectively, and the ideal distances were uniformly ≥17 ?. The limit of detection of the double-antibody sandwich and antibody-avidin sandwich modes were 1.8 and 0.8 pg/mL (calculated using AOZ), respectively. Compared with competitive ELISA, the developed ELISA method achieved a 25-fold improvement in sensitivity. The average recovery rate and average relative standard deviation were 73%–85% and 9.0%, respectively.

Key words: Furazolidone, micromolecule, sandwich immunoassay

中图分类号:

R392-33

张世伟冯荣虎吴会玲郭继平周迎春王炳志杨星星汤璐. 基于衍生化3-氨基-2-恶唑烷酮的夹心酶联免疫分析法研究[J]. 食品科学.

feng ronghu. A sandwich immunosorbent assay for the small molecule compound 3-Amino-2-oxazolidinone based on derivatization[J]. FOOD SCIENCE.

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