運用16S核醣DNA/RNA探針結合金奈米粒子之側流免疫分析檢測沙門氏菌

Salmonella detection using 16S ribosomal DNA/RNA probe-gold nanoparticles and lateral flow immunoassay

劉正哲(Cheng-Che Liu)

指導教授 ： 侯劭毅

國立臺北科技大學/工程學院/工程科技研究所/博士(2013年)

https://doi.org/10.6841/NTUT.2013.00670

摘要

本實驗開發一種靈敏、簡單且快速，可用於檢測沙門氏菌的側流免疫分析。其原理是將奈米金粒子接合上一股可互補沙門氏菌16S 核醣DNA與核醣RNA的核酸探針。首先在模擬目標序列的實驗中，運用合成之單股DNA為目標模板進行前測試驗，檢出極限值為5 fmol。本分析法實際應用於培養之沙門氏菌樣本測試，可快速於30分鐘內測得107菌數所萃取之核酸。若輔以銀還原放大訊號，則檢測極限可達104菌數，低於文獻所報導人類攝入沙門氏菌產生臨床癥候之感染菌量—105菌數。此外，相較於其他9株腸內菌（測試結果），本實驗所使用之探針對沙門氏菌確具專一性。故此分析法不僅可應用於微生物檢測，對於食品安全或臨床診斷亦為有效工具，且其兼具靈敏度、專一性及方便操作等優點，適合提供發展中國家之大規模篩選使用。

關鍵字

沙門氏菌檢測 ； 16S核醣核酸 ； 金奈米粒子 ； 側流免疫分析

並列摘要

We develop an ultrasensitive, simple, and fast lateral flow immunoassay for Salmonella detection using gold nanoparticles conjugated with a DNA probe, which is complementary to the 16S ribosomal RNA and DNA of Salmonella. The detection limit is 5 fmol for the synthetic single-stranded DNA. For Salmonella cultured samples, the nucleic acids from 107 bacteria were rapidly detected in 30 minutes. After silver enhancement, the detection limit was as low as 104 cells which is lower than 105 bacteria cells, the human infective dose of food-borne Salmonella. Furthermore, the probes used in this study are specific to Salmonella compared to 9 other species of Enterobacteriaceae. This approach would be useful for microbial detection in regards to food safety or clinical diagnosis. It is also suitable for large-scale screening in developing countries because it is sensitive, specific and convenient.

並列關鍵字

Salmonella detection ； 16S rRNA ； gold nanoparticles ； lateral flow immunoassay

參考文獻

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