Self-Digitization Microfluidic Device for Digital PCR

描述（由申请人提供）：SBIR一期项目将确定将自我数字化（SD）微流控技术作为具有商业竞争力的核酸定量（NAQ）平台的可行性。核酸（包括DNA和RNA）的定量用于包括一般研究，生物医学研究和临床诊断在内的各个领域，在诸如测量基因表达水平，疾病诊断（包括病原体/病毒载量监测），癌症诊断等领域具有特定应用。自数字化是一种简单、强大的技术，它有效地利用通道和井的网络，自发地将样品划分到一系列预定义的井体中。该平台非常适合数字PCR （dPCR）应用。数字PCR的工作原理是将样品分成数千甚至数百万个单独的体积，每个体积可能包含也可能不包含目标DNA。只有含有目标DNA的体积才能给出阳性信号，从而产生“数字”是/否信号，由此泊松统计可以直接确定样品浓度。数字PCR直接提供绝对定量，对变异具有鲁棒性

DESCRIPTION (provided by applicant): This SBIR Phase I project will determine the feasibility of using Self-Digitization (SD) microfluidic technology as a commercially competitive nucleic acid quantification (NAQ) platform. Quantification of nucleic acids (both DNA and RNA) is utilized in various fields including general research, biomedical research, and clinical diagnostics, with specific applications in areas such as measuring gene expression levels, disease diagnostics including pathogen/viral load monitoring, cancer diagnostics, and more. Self-Digitization is a simple, robust technology that efficiently uses a network of channels and wells to spontaneously partition samples into an array of predefined well volumes. This platform is ideally suited for digital PCR (dPCR) applications. Digital PCR works by partitioning samples into thousands or even millions of individual volumes where each volume may or may not contain target DNA. Only volumes with target DNA give a positive signal resulting in a "digital" yes/no signal, from which Poisson statistics can then give direct determination of sample concentrations. Digital PCR directly provides absolute quantification, is robust against variations in reaction efficiency, is incredibly sensitive and has nearly unlimited resolution capabilities, making it technically superior to the current NAQ "gold standard" of qPCR. However, no commercial dPCR system is currently able to be economically competitive with qPCR. The objective of this proposal is to show the technical feasibility of a new centrifugal SD filling method within an optical disc (OD) style platform. The simplicity and flexibility of the SD system, its implementation within a method that enables parallelization and high-throughput experimentation, and the adaptation of existing commercialized components to achieve a complete platform will result in a cost-competitive product that will surpass existing commercial dPCR platforms, while also directly competing with qPCR systems. The approach will first validate the technical aspects of the platform by developing a thorough understanding of the loading/digitization mechanism, optimizing design features to maximize performance within a closed system, and to develop complete designs to meet needs in specific application categories. This will initially be carried out in scaled down versions to maximize iteration efficiency and then expanded to full prototype scale. Validation of this approach would then lead to future production of a mass producible device product and complete instrumentation setup. The product would be immediately useful for general biomedical research applications, and the workflow and nature of the closed system would also make it amenable to future clinical applications in a variety of medical fields.