Colorimetric detection of loop-mediated isothermal amplification reaction by using hydroxy naphthol blue
Colorimetric detection of loop-mediated isothermal amplification reaction by using hydroxy naphthol blue
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DOI：
10.2144/000113072
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发表时间：
2009-03-01
期刊：
影响因子：
2.7
通讯作者：
Hanaki, Ken-Ichi
中科院分区：
文献类型：
--
作者：
Goto, Motoki;Honda, Eiichi;Hanaki, Ken-Ichi
环介导等温扩增（LAMP），一种新的基因扩增方法，能够合成大量的DNA和可见的副产物，即焦磷酸镁，没有热循环。阳性反应通过反应溶液的浑浊度或在向反应溶液中加入嵌入染料后的颜色变化来指示，但使用这种染料具有一定的局限性。羟基萘酚蓝（HNB）是一种钙的金属指示剂和碱土金属离子的比色试剂，用于LAMP反应的新比色测定。向LAMP反应溶液中预先加入120 μ M HNB不会抑制扩增效率。阳性反应由颜色从紫色变为天蓝色表示。用HNB的LAMP反应也可以在96孔微孔板中进行，并且可以用酶标仪在650 nm处测量反应。利用HNB的LAMP方法将有助于高通量的DNA和RNA检测。
Loop-mediated isothermal amplification (LAMP), a novel gene amplification method, enables the synthesis of larger amounts of both DNA and a visible byproduct-namely, magnesium pyrophosphate-without thermal cycling. A positive reaction is indicated by the turbidity of the reaction solution or the color change after adding an intercalating dye to the reaction solution, but the use of such dyes has certain limitations. Hydroxy naphthol blue (HNB), a metal indicator for calcium and a colorimetric reagent for alkaline earth metal ions, was used for a new colorimetric assay of the LAMP reaction. Preaddition of 120 mu M HNB to the LAMP reaction solution did not inhibit amplification efficiency. A positive reaction is indicated by a color change from violet to sky blue. The LAMP reaction with HNB could also be carried out in a 96-well microplate, and the reaction could be measured at 650 nm with a microplate reader. The colorimetric LAMP method using HNB would be helpful for high-throughput DNA and RNA detection.