Gold Nanoparticle-Based Enzyme-Linked Antibody-Aptamer Sandwich Assay for Detection of Salmonella Typhimurium
Gold Nanoparticle-Based Enzyme-Linked Antibody-Aptamer Sandwich Assay for Detection of Salmonella Typhimurium
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DOI：
10.1021/am5045828
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发表时间：
2014-10-08
影响因子：
9.5
通讯作者：
Lu, Jianxin
中科院分区：
文献类型：
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作者：
Wu, Wenhe;Li, Jun;Lu, Jianxin
酶联免疫吸附试验（ELISA）为快速检测肠道沙门氏菌血清型鼠伤寒沙门氏菌（STM）提供了一种便捷的方法，对食源性致病菌的快速诊断具有重要意义。然而，传统的酶联免疫吸附试验受抗体-抗原免疫反应的限制，灵敏度差，样品前处理繁琐。因此，新型酶联免疫吸附试验的开发仍然具有挑战性。为此，我们设计了一种基于金纳米粒子的酶联抗体适体夹心（nano-ELAAS）快速、灵敏、定量、高特异性检测STM的综合策略。用核酸适配体修饰的磁性颗粒从样品中捕获并预浓缩STM，然后与检测抗体结合。然后利用携带大量报告抗体和辣根过氧化物酶分子的纳米探针进行比色信号放大。在优化的反应条件下，纳米elaas法的定量检测范围为1 × 10(3) ~ 1 × 10(8) CFU mL(-1)，检测限为1 × 10(3) CFU mL(-1)，在含有其他细菌的较高浓度样品中，STM的选择性为bb10倍，检测时间小于3 h。此外，与两种商用酶标记抗体信号报告器相比，所开发的纳米探针在检测范围和/或灵敏度方面均有提高。最后，纳米elaas方法被证明在牛奶样品中工作良好，牛奶样品是STM污染的常见来源。
Enzyme-linked immunosorbent assay (ELISA) provides a convenient means for the detection of Salmonella enterica serovar Typhimurium (STM), which is important for rapid diagnosis of foodborne pathogens. However, conventional ELISA is limited by antibody-antigen immunoreactions and suffers from poor sensitivity and tedious sample pretreatment. Therefore, development of novel ELISA remains challenging. Herein, we designed a comprehensive strategy for rapid, sensitive, and quantitative detection of STM with high specificity by gold nanoparticle-based enzyme-linked antibody-aptamer sandwich (nano-ELAAS) method. STM was captured and preconcentrated from samples with aptamer-modified magnetic particles, followed by binding with detector antibodies. Then nanoprobes carrying a large amount of reporter antibodies and horseradish peroxidase molecules were used for colorimetric signal amplification. Under the optimized reaction conditions, the nano-ELAAS assay had a quantitative detection range from 1 X 10(3) to 1 X 10(8) CFU mL(-1), a limit of detection of 1 X 10(3) CFU mL(-1), and a selectivity of >10-fold for STM in samples containing other bacteria at higher concentration with an assay time less than 3 h. In addition, the developed nanoprobes were improved in terms of detection range and/or sensitivity when compared with two commercial enzyme-labeled antibody signal reporters. Finally, the nano-ELAAS method was demonstrated to work well in milk samples, a common source of STM contamination.