A fully integrated microfluidic genetic analysis system with sample-in-answer-out capability
A fully integrated microfluidic genetic analysis system with sample-in-answer-out capability
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DOI：
10.1073/pnas.0604663103
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发表时间：
2006-12-19
影响因子：
11.1
通讯作者：
Landers, James P.
中科院分区：
文献类型：
--
作者：
Easley, Christopher J.;Karlinsey, James M.;Landers, James P.
我们描述了一种微流体遗传分析系统，该系统代表了一种先前未描述的集成微流体装置，能够接受全血作为粗生物样本，并最终生成遗传图谱。加载样品后，玻璃微流控遗传分析系统装置会进行片上 DNA 纯化和基于 PCR 的扩增，然后进行分离和检测，从而可以在 30 分钟内对微升样品进行传染性病原体筛查，并获得样品输入-应答输出结果。单个注射泵将样品/试剂输送至芯片以从生物样品中纯化核酸。弹性膜阀门隔离装置的每个不同功能区域，并与阻力流一起将纯化的 DNA 和 PCR 试剂从提取域引导到 550-nl 室中，以进行快速靶序列 PCR 扩增。基于压力的重复注射纳升等份扩增子（以及 DNA 大小标准）允许电泳分离和检测，以提供 DNA 片段大小信息。所得到的遗传谱证实了来自活体无症状感染小鼠的 750 nl 全血中存在炭疽杆菌（炭疽），以及来自疑似百日咳患者的 1 μl 鼻吸出物中存在百日咳博德特氏菌。
We describe a microfluidic genetic analysis system that represents a previously undescribed integrated microfluidic device capable of accepting whole blood as a crude biological sample with the endpoint generation of a genetic profile. Upon loading the sample, the glass microfluidic genetic analysis system device carries out on-chip DNA purification and PCR-based amplification, followed by separation and detection in a manner that allows for microliter samples to be screened for infectious pathogens with sample-in-answer-out results in < 30 min. A single syringe pump delivers sample/reagents to the chip for nucleic acid purification from a biological sample. Elastomeric membrane valving isolates each distinct functional region of the device and, together with resistive flow, directs purified DNA and PCR reagents from the extraction domain into a 550-nl chamber for rapid target sequence PCR amplification. Repeated pressure-based injections of nanoliter ali-quots of amplicon (along with the DNA sizing standard) allow electrophoretic separation and detection to provide DNA fragment size information. The presence of Bacillus anthracis (anthrax) in 750 nl of whole blood from living asymptomatic infected mice and of Bordetella pertussis in 1 mu l of nasal aspirate from a patient suspected of having whooping cough are confirmed by the resultant genetic profile.