基于EWOD的单片集成多体积数字PCR研究

The digital polymerase chain reaction (dPCR) has shown great advantages over conventional PCR in direct absolute quantification, low-concentration template detection, nuanced difference detection and rare mutation detection. It is an imperative hot research for developing a dPCR system which is miniaturized, portable and automatic for practical applications. By far the systematic relationships between active controls of micro-droplets, thermal conduction between micro-interfaces and integration of chips have not been researched intensively in dPCR, which urgently need to be clarified and integrated..An innovative EWOD dPCR device based on MV-dPCR (Multivolume dPCR) principle is proposed, with the remarkable advantages of generating low-volume, multi-volume droplets array in simultaneously, integrating electrode multiplexing technology to achieve monolithic, automaticity droplets control, detection and analysis by smart phone. The main innovation points of the research are as follows: 1. An EWOD based integrated single-chip MV-dPCR system is proposed; 2. The mechanism of nL/pL level multi-droplet generation is proposed; 3. The thermal conduction law between multi-phase heterogeneous interfaces of high speed micro-droplets is explored..The innovative EWOD based portable dPCR system is a new type of dPCR system with independent intellectual property, which will enhance the international competitiveness of China in the research field of molecular diagnostic instruments.

数字聚合酶链式反应（dPCR）在直接绝对定量、低浓度模板检测、细微差异检测、稀有变异检测等方面相比传统PCR有巨大的优势，发展微小型/便携和自动化操作的dPCR系统是热点之一。dPCR技术在微液滴主动控制、微观界面热传导、芯片集成化方面还没有被系统性深入研究，三者之间的深层次关系亟待需要阐明和整合。.本项目提出了一种基于介电润湿（EWOD）原理的多体积液滴dPCR（MV-dPCR），充分发挥EWOD并行产生低通量多体积阵列液滴的优势，同时利用电极复用技术实现单片集成的dPCR，并利用智能手机实现液滴控制、检测和分析。创新点：1、提出了EWOD式单芯片集成的MV-dPCR系统；2、提出了nL/pL级多液滴产生机理；3、探究了运动中微液滴在多相界面的热传导规律。.本项目提出的基于EWOD的便携式MV-dPCR具有自主知识产权，是一种新型的dPCR系统，将增强我国在分子诊断仪器领域的国际竞争力。

数字聚合酶链式反应（dPCR）在直接绝对定量、低浓度模板检测、细微差异检测、稀有变异检测等方面相比传统PCR有巨大的优势，发展微小型/便携和自动化操作的dPCR系统是热点之一。.项目实现了两种面向dPCR应用、利用电极复用技术实现单片集成加热器芯片，利用智能手机和机器学习实现液滴控制、检测和分析。项目充分发挥EWOD力作用下的仿生多体积微液滴阵列芯片的优势，采用一步法实现几个数量级的大面积pL微液滴生成，在dPCR中的应用表明：芯片灵敏度和动态监测增加2个数量级，检测上限增加一个数量级。项目在新型EWOD器件、智能识别微液滴、pL级大面积微液滴生成及多体积dPCR应用几方面具有创新。.项目提出的基于EWOD的创新dPCR系统具有完全自主知识产权，将会增强我国在分子诊断仪器研究领域的国际竞争力。.共发表期刊论文19篇（其中国际期刊18篇），IEEE国际会议论文5篇。申请发明专利5项，授权共8项（其中1项为项目执行期内申请并获授权）。培养博士毕业生6人（其中3人毕业论文为优秀）和硕士研究生2人。