Shanghai Key Lab of Modern Optical System, School of Optical-Electrical and Computer Engineering, University of Shanghai for Science and Technology, Shanghai 200093, China.
Shanghai Turtle Technology Limited, Shanghai 200439, China.
Lab Chip. 2021 Sep 28;21(19):3742-3747. doi: 10.1039/d1lc00532d.
Droplet digital polymerase chain reaction (ddPCR) is an effective technique, with unparalleled sensitivity, for the absolute quantification of target nucleic acids. However, current commercial ddPCR devices for detecting the gene chip are time consuming due to complex image stitching. To address this issue, we propose a universal concentration determination system and realize one-time gene chip imaging with high resolution. All the functional units are controlled by self-developed software using the PyQt5 module in Python. Without stitching technology, images of the ddPCR chip (28 mm × 18 mm) containing 20 000 independent 0.81 nL micro chambers can be obtained in less than 15 seconds, which saves about 165 seconds. A white laser light source (2 mW cm) was employed as a substitute for the mercury lamp. Its wavelength matches well with typical fluorescent dyes (, HEX, ROX and Cy5), and thus it can strengthen the fluorescence intensity for weak signals. The results also demonstrated that the correlation coefficient for the measured concentration and theoretical value was above 99%, by testing the ddPCR products with COVID-19 virus. Such a system can greatly reduce the time required for image acquisition and DNA concentration determination, and thus is able to speed up the lab-to-application process for ddPCR technology.
微滴式数字聚合酶链式反应（ddPCR）是一种有效的技术，具有无与伦比的灵敏度，可用于绝对定量目标核酸。然而，目前用于检测基因芯片的商用 ddPCR 设备由于复杂的图像拼接而耗时。为了解决这个问题，我们提出了一种通用的浓度测定系统，并实现了一次成像的高分辨率基因芯片。所有的功能单元都由我们自主开发的软件控制，使用 Python 中的 PyQt5 模块。不使用拼接技术，在不到 15 秒的时间内就可以获取包含 20000 个独立的 0.81 nL 微腔的 ddPCR 芯片（28mm×18mm）的图像，节省了约 165 秒。我们使用 2mWcm 的白色激光光源替代汞灯。其波长与典型的荧光染料（FAM、HEX、ROX 和 Cy5）很好地匹配，因此可以增强弱信号的荧光强度。通过检测 COVID-19 病毒的 ddPCR 产物，结果表明测量浓度与理论值的相关系数大于 99%。该系统可以大大减少图像采集和 DNA 浓度测定所需的时间，从而加快 ddPCR 技术的实验室应用进程。
