Digital PCR is a powerful method for absolute nucleic acid quantification and is widely used in the absolute quantification of viral copy numbers, tumor marker detection, and prenatal diagnosis. However, for most of the existing droplet-based dPCR systems, the droplet generation, PCR reaction, and droplet detection are performed separately using different instruments. Making digital PCR both easy to use and practical by integrating the qPCR workflow into a superior all-in-one walkaway solution is one of the core ideas. A new innovative and integrated digital droplet PCR platform was developed that utilizes cutting-edge microfluidics to integrate dPCR workflows onto a single consumable chip. This makes previously complex workflows fast and simple; the whole process of droplet generation, PCR amplification, and droplet detection is completed on one chip, which meets the clinical requirement of “sample in, result out”. It provides high multiplexing capabilities and strong sensitivity while all measurements were within the 95% confidence interval. This study is the first validation of the DropXpert S6 system and focuses primarily on verifying its reliability, repeatability, and consistency. In addition, the accuracy, detection limit, linearity, and precision of the system were evaluated after sample collection. Among them, the accuracy assessment by calculating the absolute bias of each target gene yielded a range from −0.1 to 0.08, all within ±0.5 logarithmic orders of magnitude; the LOB for the assay was set at 0, and the LoD value calculated using probit curves is MR4.7 (0.002%); the linearity evaluation showed that the R² value of the BCR-ABL was 0.9996, and the R² value of the ABL metrics calculated using the ERM standard was 0.9999; and the precision evaluation showed that all samples had a CV of less than 4% for intra-day, inter-day, and inter-instrument variation. The CV of inter-batch variation was less than 7%. The total CV was less than 5%. The results of the study demonstrate that dd-PCR can be applied to molecular detection and the clinical evaluation of CML patients and provide more precise personal treatment guidance, and its reproducibility predicts the future development of a wide range of clinical applications.

中文翻译：

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DropXpert S6 系统诊断慢性粒细胞白血病的分析验证


数字PCR是一种强大的核酸绝对定量方法，广泛应用于病毒拷贝数绝对定量、肿瘤标志物检测和产前诊断等领域。然而，对于大多数现有的基于液滴的 dPCR 系统，液滴生成、PCR 反应和液滴检测是使用不同的仪器分别进行的。通过将 qPCR 工作流程集成到卓越的一体式解决方案中，使数字 PCR 既易于使用又实用，这是其核心理念之一。开发了一种新的创新集成数字液滴 PCR 平台，该平台利用尖端的微流控技术将 dPCR 工作流程集成到单个耗材芯片上。这使得以前复杂的工作流程变得快速而简单；液滴生成、PCR扩增、液滴检测全过程在一块芯片上完成，满足“进样、出结果”的临床要求。它提供高复用能力和强大的灵敏度，同时所有测量都在 95% 置信区间内。本研究是对 DropXpert S6 系统的首次验证，主要侧重于验证其可靠性、可重复性和一致性。此外，样品采集后还对系统的准确度、检出限、线性度和精密度进行了评估。其中，通过计算每个目标基因的绝对偏差进行准确度评估，得出的范围为-0.1至0.08，均在±0.5个对数数量级之内；检测的 LOB 设置为 0，使用概率曲线计算的 LoD 值为 MR4.7 (0.002%)；线性评估结果显示，BCR-ABL的R ² 值为0.9996，使用ERM标准计算的ABL指标的R ² 值为0。9999；精密度评估显示，所有样品的日内、日间、仪器间变异CV均小于4%。批次间变异CV小于7%。总CV小于5%。研究结果表明，dd-PCR可应用于CML患者的分子检测和临床评估，并提供更精准的个体化治疗指导，其重现性预示着未来广泛临床应用的发展。