3D printing and milling a real-time PCR device for infectious disease diagnostics.
3D printing and milling a real-time PCR device for infectious disease diagnostics.
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DOI：
10.1371/journal.pone.0179133
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Kim BN
中科院分区：
文献类型：
--
作者：
Mulberry G;White KA;Vaidya M;Sugaya K;Kim BN
利用定量聚合酶链式反应(QPCR)诊断传染病，可为确定感染、病原体的菌株或类型以及感染程度提供决定性的结果。然而，由于涉及的仪器设备成本高，维护复杂，在现场很少使用它来进行快速故障诊断。为了提供比现有定量聚合酶链式反应装置更高水平的可获得性，探索了一组3D制造方法作为制造低成本和便携式的定量聚合酶链式反应装置的可能选项。这种方法的主要优势是能够将设计文件的数字格式上传到互联网上，以便广泛分发，这样无论在哪里，人们都可以简单地下载并输入他们的3D打印机，以便快速制造。材料和设计经过精心选择，以最大限度地减少依赖于先进制造工艺的定制部件的数量，这降低了可访问性。用含有不同浓度慢病毒的20μL样本对所提出的3D制造的定量聚合酶链式反应装置进行了测试，慢病毒与艾滋病毒的类型相同。反转录步骤是设备操作的一部分，该步骤发生在qPCR步骤之前，以将慢病毒的目标RNA反转录为互补DNA(CDNA)。紧随其后的是定量聚合酶链式反应，它在聚合酶链式反应扩增过程中对样品中的目标序列分子进行定量。热控和时间协调荧光读数的整个过程由闭环反馈和微控制器实现自动化。该装置是便携的，电池供电，尺寸为12×7×6厘米3，质量仅为214克。通过在线上传和共享设计文件，所提出的低成本定量聚合酶链式反应装置可能更容易获得对各种传染病，如艾滋病毒和疟疾的强大诊断方案。
Diagnosing infectious diseases using quantitative polymerase chain reaction (qPCR) offers a conclusive result in determining the infection, the strain or type of pathogen, and the level of infection. However, due to the high-cost instrumentation involved and the complexity in maintenance, it is rarely used in the field to make a quick turnaround diagnosis. In order to provide a higher level of accessibility than current qPCR devices, a set of 3D manufacturing methods is explored as a possible option to fabricate a low-cost and portable qPCR device. The key advantage of this approach is the ability to upload the digital format of the design files on the internet for wide distribution so that people at any location can simply download and feed into their 3D printers for quick manufacturing. The material and design are carefully selected to minimize the number of custom parts that depend on advanced manufacturing processes which lower accessibility. The presented 3D manufactured qPCR device is tested with 20-μL samples that contain various concentrations of lentivirus, the same type as HIV. A reverse-transcription step is a part of the device’s operation, which takes place prior to the qPCR step to reverse transcribe the target RNA from the lentivirus into complementary DNA (cDNA). This is immediately followed by qPCR which quantifies the target sequence molecules in the sample during the PCR amplification process. The entire process of thermal control and time-coordinated fluorescence reading is automated by closed-loop feedback and a microcontroller. The resulting device is portable and battery-operated, with a size of 12 × 7 × 6 cm3 and mass of only 214 g. By uploading and sharing the design files online, the presented low-cost qPCR device may provide easier access to a robust diagnosis protocol for various infectious diseases, such as HIV and malaria.