An isothermal amplification reactor with an integrated isolation membrane for point-of-care detection of infectious diseases.
An isothermal amplification reactor with an integrated isolation membrane for point-of-care detection of infectious diseases.
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DOI：
10.1039/c1an00007a
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发表时间：
2011-05-21
期刊：
影响因子：
--
通讯作者：
Bau HH
中科院分区：
文献类型：
--
作者：
Liu C;Geva E;Mauk M;Qiu X;Abrams WR;Malamud D;Curtis K;Owen SM;Bau HH
设计、构建并测试了一种用于检测从病原体中提取的核酸的简单、即时、廉价、一次性试剂盒。该试剂盒利用单个反应室进行核酸等温扩增。该腔室配备有用于分离、浓缩和纯化DNA和/或RNA的集成的流通式Flinders Technology Associates（Whatman FTA®）膜。被膜捕获的核酸直接用作扩增模板而无需洗脱，从而简化了盒的流动控制。FTA膜还起到另一个关键作用，即能够去除显著降低检测灵敏度的抑制剂。热控制装置配备有卡匣外部的薄膜加热器。扩增过程用便携式紧凑型荧光读数器进行真实的实时监测。通过检测口腔液中HIV-1的存在证明了集成单腔盒的实用性。逆转录HIV RNA并进行环介导等温扩增（LAMP）。证明了小于10个HIV颗粒的检测限。这种盒式磁带特别适用于资金和训练有素的人员短缺的资源贫乏地区。该盒可以容易地被修改以检测与唾液、尿液和其他体液以及水和食物中携带的其他病原体相关的核酸。
A simple, point of care, inexpensive, disposable cassette for the detection of nucleic acids extracted from pathogens was designed, constructed, and tested. The cassette utilizes a single reaction chamber for isothermal amplification of nucleic acids. The chamber is equipped with an integrated, flow-through, Flinders Technology Associates (Whatman FTA®) membrane for the isolation, concentration, and purification of DNA and/or RNA. The nucleic acids captured by the membrane are used directly as templates for amplification without elution, thus simplifying the cassette’s flow control. The FTA membrane also serves another critical role—enabling the removal of inhibitors that dramatically reduce detection sensitivity. Thermal control is provided with a thin film heater external to the cassette. The amplification process was monitored in real time with a portable, compact fluorescent reader. The utility of the integrated, single-chamber cassette was demonstrated by detecting the presence of HIV-1 in oral fluids. The HIV RNA was reverse transcribed and subjected to loop-mediated, isothermal amplification (LAMP). A detection limit of less than 10 HIV particles was demonstrated. The cassette is particularly suitable for resource poor regions, where funds and trained personnel are in short supply. The cassette can be readily modified to detect nucleic acids associated with other pathogens borne in saliva, urine, and other body fluids as well as in water and food.
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影响因子：
12.7
作者：
Curtis, Kelly A.;Rudolph, Donna L.;Owen, Michele
通讯作者：
Owen, Michele
影响因子：
4.2
作者：
Kim, Jitae;Mauk, Michael;Bau, Haim H.
通讯作者：
Bau, Haim H.
影响因子：
6.1
作者：
Lagally, ET;Emrich, CA;Mathies, RA
通讯作者：
Mathies, RA
DOI：
10.1073/pnas.0607254104
发表时间：
2007-03-27
影响因子：
11.1
作者：
Herr, Amy E.;Hatch, Anson V.;Singh, Anup K.
通讯作者：
Singh, Anup K.
DOI：
10.1073/pnas.0604663103
发表时间：
2006-12-19
影响因子：
11.1
作者：
Easley, Christopher J.;Karlinsey, James M.;Landers, James P.
通讯作者：
Landers, James P.