用于全自动数字 PCR 图像处理的图像到答案算法,Lab on a Chip

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用于全自动数字 PCR 图像处理的图像到答案算法
Lab on a Chip ( IF 6.1 ) Pub Date : 2022-02-24 , DOI: 10.1039/d1lc01175h
Zhiqiang Yan ₁ , Haoqing Zhang ₁ , Xinlu Wang ₁ , Martina Gaňová ₂ , Tomáš Lednický ₂ , Hanliang Zhu ₁ , Xiaocheng Liu ₁ , Marie Korabečná ₃ , Honglong Chang ₁ , Pavel Neužil ₁

Affiliation

数字聚合酶链式反应 (dPCR) 是 PCR 技术不可替代的变体，因为它具有绝对定量和检测临床样本中稀有脱氧核糖核酸 (DNA) 序列的能力。图像处理方法，包括微室定位和荧光分析，确定 dPCR 结果的可靠性。然而，典型的方法对芯片结构、芯片填充和光强均匀性要求很高。这项研究开发了一种图像到答案算法，具有单荧光图像捕获和已知的图像相关错误消除。我们应用霍夫变换来识别 dPCR 芯片图像中的分区，应用 2D 傅里叶变换来旋转图像，并应用 3D 投影变换来定位和校正所有分区的位置。然后我们计算每个分区的平均荧光幅度并生成图像的 3D 荧光强度分布图。我们随后根据图谱校正了分区之间的荧光不均匀性，并获得了分区荧光强度的统计结果。我们使用目标 DNA 的不同内容验证了所提出的算法。所提出的算法与 dPCR 芯片结构损坏和光强不均匀性无关。它还提供了一种可靠的替代方法来分析基于芯片的 dPCR 系统的结果。我们随后根据图谱校正了分区之间的荧光不均匀性，并获得了分区荧光强度的统计结果。我们使用目标 DNA 的不同内容验证了所提出的算法。所提出的算法与 dPCR 芯片结构损坏和光强不均匀性无关。它还提供了一种可靠的替代方法来分析基于芯片的 dPCR 系统的结果。我们随后根据图谱校正了分区之间的荧光不均匀性，并获得了分区荧光强度的统计结果。我们使用目标 DNA 的不同内容验证了所提出的算法。所提出的算法与 dPCR 芯片结构损坏和光强不均匀性无关。它还提供了一种可靠的替代方法来分析基于芯片的 dPCR 系统的结果。

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An image-to-answer algorithm for fully automated digital PCR image processing

The digital polymerase chain reaction (dPCR) is an irreplaceable variant of PCR techniques due to its capacity for absolute quantification and detection of rare deoxyribonucleic acid (DNA) sequences in clinical samples. Image processing methods, including micro-chamber positioning and fluorescence analysis, determine the reliability of the dPCR results. However, typical methods demand high requirements for the chip structure, chip filling, and light intensity uniformity. This research developed an image-to-answer algorithm with single fluorescence image capture and known image-related error removal. We applied the Hough transform to identify partitions in the images of dPCR chips, the 2D Fourier transform to rotate the image, and the 3D projection transformation to locate and correct the positions of all partitions. We then calculated each partition's average fluorescence amplitudes and generated a 3D fluorescence intensity distribution map of the image. We subsequently corrected the fluorescence non-uniformity between partitions based on the map and achieved statistical results of partition fluorescence intensities. We validated the proposed algorithms using different contents of the target DNA. The proposed algorithm is independent of the dPCR chip structure damage and light intensity non-uniformity. It also provides a reliable alternative to analyze the results of chip-based dPCR systems.

更新日期：2022-02-24

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