RT-LAMP检测甲型H1N1病毒核酸几种结果判定方法

Comparable Study on Determination Methods for Detection of Influenza A(H1N1) Virus Nucleic Acid by RT-LAMP

目的比较RT-LAMP检测甲型H1N1流感病毒核酸几种不同的结果判定方法的差异,优化检测方法。方法利用已知的阳性病毒样本,对比电泳、直接观察、加入SYBR GREENⅠ核酸染料和优化后的预染核酸染料和的检测灵敏度。结果电泳检测的灵敏度最高,加入SYBR GREENⅠ染料的灵敏度略低,而直接的肉眼观察灵敏度低约2个数量级。加入优化后的预染染料其检测灵敏度与加入SYBR GREENⅠ核酸染料相当。结论电泳法检测的灵敏度最高,可通过加入优化后的预染染料提高目测判定反应结果的灵敏度,增强反应的特异性,并可降低气溶胶污染。

Objective Four different determination process were compared to optimize method of detection for influenza A （H1NI） virus nucleic acid by RT - LAMP. Methods With the confirmed positive samples, the detection sensitivity of specific nucleic acid by direct visual observation, adding dyes after reaction, electrophoresis, and adding optimized dye in reaction mixture before reaction were analyzed. Results Electrophoresis detection was the most sensitive among four detection methods, and the sensitivity of adding SYBR GREEN I dye after reaction was inferior to it. Howerer, the sensitivity of direct visual observation was 100 fold lower than that of electrophoresis detection. The sensitivity of adding optimized dye in reaction mixture before reaction was comparable to that of adding dyes after reaction. Conclusion Electrophoresis is the most sensitive determination method of LAMP. The visual observation sensitivity of RT - LAMP can be improved by adding the optimized dye before reaction, while the specificity can be enhanced, and the aerosol contamination can also be diminished.