An integrated microfluidic platform for sensitive and rapid detection of biological toxins
An integrated microfluidic platform for sensitive and rapid detection of biological toxins
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DOI：
10.1039/b815152k
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发表时间：
2008-01-01
期刊：
影响因子：
6.1
通讯作者：
Singh, Anup K.
中科院分区：
文献类型：
--
作者：
Meagher, Robert J.;Hatch, Anson V.;Singh, Anup K.
为了设计用于检测体液中的生物毒素的便携式诊断装置，我们已经开发了基于微流控芯片的免疫测定，其快速（< 20分钟），需要最小的样品体积（< 10 μ L），并且具有可观的灵敏度和动态范围（μ M-pM）。微流控芯片正在与微型电子器件、光学元件、流体处理组件和数据采集软件集成，以开发一种便携式、独立的设备。该器械预期用于在意外或故意暴露/中毒于生物毒素的情况下进行快速、即时（以及未来的事件点）检测。毒素和潜在的宿主反应标志物的检测是使用微流控电泳免疫测定与样品预浓缩和混合的分析物与荧光标记的抗体集成。通过在芯片的样品加载区域中光聚合具有类似于10 kDa的MW截止的薄的纳米多孔膜来实现预浓缩。具有较大孔的聚合物凝胶位于尺寸排阻膜附近，以进行抗体-分析物复合物和过量抗体的电泳分离。使用灵敏的激光诱导荧光检测来测量结合的和未结合的免疫复合物的比率提供了样品中分析物的定量。我们已经证明了生物毒素蓖麻毒素，滋贺毒素I，和葡萄球菌肠毒素B（SE B）的电泳免疫测定。在芯片外混合且无样品预浓缩的情况下，SEB的检测限（LOD）为300 pM，滋贺毒素I的检测限为500 pM，蓖麻毒素的检测限为20 nM。在10分钟芯片上预浓缩的情况下，SEB的LOD < 10 pM。正在开发的便携式设备很容易适用于检测许多其他疾病的蛋白质生物标志物，并旨在代表下一代诊断设备，能够同时快速和定量测量多种分析物。
Towards designing a portable diagnostic device for detecting biological toxins in bodily fluids, we have developed microfluidic chip-based immunoassays that are rapid (< 20 minutes), require minimal sample volume (< 10 mu L) and have appreciable sensitivity and dynamic range (mu M-pM). The microfluidic chip is being integrated with miniaturized electronics, optical elements, fluid-handling components, and data acquisition software to develop a portable, self-contained device. The device is intended for rapid, point-of-care (and, in future, point-of-incident) testing in case of an accidental or intentional exposure/intoxication to biotoxins. Detection of toxins and potential host-response markers is performed using microfluidic electrophoretic immunoassays integrated with sample preconcentration and mixing of analytes with fluorescently labeled antibodies. Preconcentration is enabled by photopolymerizing a thin, nanoporous membrane with a MW cut-off of similar to 10 kDa in the sample loading region of the chip. Polymeric gels with larger pores are located adjacent to the size exclusion membrane to perform electrophoretic separation of antibody-analyte complex and excess antibody. Measurement of the ratio of bound and unbound immune-complex using sensitive laser-induced fluorescence detection provides quantitation of analyte in the sample. We have demonstrated electrophoretic immunoassays for the biotoxins ricin, Shiga toxin I, and Staphylococcal enterotoxin B (SEB). With off-chip mixing and no sample preconcentration, the limits of detection (LOD) were 300 pM for SEB, 500 pM for Shiga toxin I, and 20 nM for ricin. With a 10 min on-chip preconcentration, the LOD for SEB is < 10 pM. The portable device being developed is readily applicable to detection of proteinaceous biomarkers of many other diseases and is intended to represent the next-generation diagnostic devices capable of rapid and quantitative measurements of multiple analytes simultaneously.