Computational Bioimaging and Bioinformatics, Institut de Biologie de l'Ecole Normale Supérieure, PSL University, 46 Rue d'Ulm, 75005, Paris, France.
Biophenics Laboratory, Department of Translational Research, Cell and Tissue Imaging Facility (PICT-IBiSA), Institut Curie, PSL Research University, 26 Rue d'Ulm, 75005, Paris, France.
Sci Rep. 2023 Dec 18;13(1):22599. doi: 10.1038/s41598-023-49554-8.
High content screening (HCS) is a technology that automates cell biology experiments at large scale. A High Content Screen produces a high amount of microscopy images of cells under many conditions and requires that a dedicated image and data analysis workflow be designed for each assay to select hits. This heavy data analytic step remains challenging and has been recognized as one of the burdens hindering the adoption of HCS. In this work we propose a solution to hit selection by using transfer learning without additional training. A pretrained residual network is employed to encode each image of a screen into a discriminant representation. The deep features obtained are then corrected to account for well plate bias and misalignment. We then propose two training-free pipelines dedicated to the two main categories of HCS for compound selection: with or without positive control. When a positive control is available, it is used alongside the negative control to compute a linear discriminant axis, thus building a classifier without training. Once all samples are projected onto this axis, the conditions that best reproduce the positive control can be selected. When no positive control is available, the Mahalanobis distance is computed from each sample to the negative control distribution. The latter provides a metric to identify the conditions that alter the negative control's cell phenotype. This metric is subsequently used to categorize hits through a clustering step. Given the lack of available ground truth in HCS, we provide a qualitative comparison of the results obtained using this approach with results obtained with handcrafted image analysis features for compounds and siRNA screens with or without control. Our results suggests that the fully automated and generic pipeline we propose offers a good alternative to handcrafted dedicated image analysis approaches. Furthermore, we demonstrate that this solution select conditions of interest that had not been identified using the primary dedicated analysis. Altogether, this approach provides a fully automated, reproducible, versatile and comprehensive alternative analysis solution for HCS encompassing compound-based or downregulation screens, with or without positive controls, without the need for training or cell detection, or the development of a dedicated image analysis workflow.
高通量筛选(HCS)是一种能够大规模自动化细胞生物学实验的技术。高通量筛选会生成大量在多种条件下的细胞显微镜图像,并且需要为每个检测设计专门的图像和数据分析工作流程,以选择命中结果。这个繁重的数据分析步骤仍然具有挑战性,并且已被认为是阻碍高通量筛选采用的一个负担。在这项工作中,我们提出了一种无需额外训练即可进行命中选择的解决方案,即使用迁移学习。使用预训练的残差网络将屏幕的每张图像编码为判别表示。然后,对所获得的深度特征进行校正,以解决板孔偏差和对准问题。然后,我们提出了两种与化合物选择相关的、适用于主要两类高通量筛选的无训练流水线:有阳性对照和无阳性对照。当有阳性对照可用时,它与阴性对照一起用于计算线性判别轴,从而在不进行训练的情况下构建分类器。一旦将所有样本都投影到该轴上,就可以选择最佳重现阳性对照的条件。当没有阳性对照时,从每个样本到阴性对照分布计算马氏距离。后者提供了一种识别改变阴性对照细胞表型的条件的度量标准。然后通过聚类步骤使用该度量标准对命中结果进行分类。由于高通量筛选中缺乏可用的真实数据,我们使用该方法获得的结果与具有或不具有对照的化合物和 siRNA 筛选的手工制作图像分析特征获得的结果进行了定性比较。我们的结果表明,我们提出的完全自动化和通用流水线是手工制作的专用图像分析方法的一个很好的替代方案。此外,我们证明该解决方案选择了使用主要专用分析方法未识别出的感兴趣条件。总的来说,该方法为高通量筛选提供了一种完全自动化、可重现、多功能和全面的替代分析解决方案,涵盖基于化合物或下调筛选、有或无阳性对照,无需培训或细胞检测,也无需开发专用的图像分析工作流程。