Miniaturized Continuous-Flow Digital PCR for Clinical-Level Serum Sample Based on the 3D Microfluidics and CMOS Imaging Device.

In recent years, the development of polymerase chain reaction (PCR) technology has focused on digital PCR, which depends on the microfluidics. Based on continuous-flow microfluidic technology, this paper designed a miniaturized digital PCR amplification system, and greatly reduced the area required for microdroplet generation and reaction. The core rod. made of polydimethylsiloxane (PDMS), was combined with the Teflon tube to form 3D microfluidics, which requires only one heating source to form the temperature difference required for gene amplification. Only two 34 g needles can form and transmit micro-droplets in a 4-fold tapered Teflon tube, which is the simplest method to generate digital PCR droplets as far as we know, which allows the microdroplet generation device to be free from dependence on expensive chips. A complementary metal oxide semiconductor (CMOS) camera was used as a detection tool to obtain fluorescence video for the entire loop area or a specified loop area. In addition, we developed a homebrew for automatic image acquisition and processing to realize the function of digital PCR. This technique realizes the analysis of clinical serum samples of hepatitis B virus (HBV) and obtained the same results as real-time quantitative PCR. This system has greatly reduced the size and cost of the entire system, while maintaining a stable response.