Dai Xiaoqing, Cao Meng, Wang Zunliang
State Key Laboratory of Digital Medical Engineering, School of Biological Science and Medical Engineering, Southeast University, Si Pai Lou 2, Nanjing 210096, China.
Biosensors (Basel). 2025 Jan 10;15(1):36. doi: 10.3390/bios15010036.
We present a cost-effective and simple multiplex nucleic acid quantification method using droplet digital PCR (ddPCR) with digital melting curve analysis (MCA). This approach eliminates the need for complex fluorescent probe design, reducing both costs and dependence on fluorescence channels. We developed a convolutional neighborhood search algorithm to correct droplet displacement during heating, ensuring precise tracking and accurate extraction of melting curves. An experimental protocol for digital MCA on the ddPCR platform was established, enabling accurate quantification of six target pathogen genes using a single fluorescence channel, with an average accuracy of 85%. Our method overcomes the multiplexing limitations of ddPCR, facilitating its application in multi-target pathogen detection.
我们提出了一种具有成本效益且简单的多重核酸定量方法，该方法使用带有数字熔解曲线分析（MCA）的微滴数字PCR（ddPCR）。这种方法无需复杂的荧光探针设计，降低了成本以及对荧光通道的依赖。我们开发了一种卷积邻域搜索算法来校正加热过程中的微滴位移，确保精确跟踪和准确提取熔解曲线。在ddPCR平台上建立了数字MCA的实验方案，能够使用单个荧光通道对六个目标病原体基因进行准确定量，平均准确率为85%。我们的方法克服了ddPCR的多重检测限制，促进了其在多目标病原体检测中的应用。
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