光学联合筛选 (OPS) 是一种可扩展的方法，用于将基于图像的表型与细胞扰动联系起来。然而，由于与扰动条形码的原位测序相关的限制，迄今为止它仅限于培养的癌细胞系中相对低复杂的表型读数。在这里，我们开发了 PerturbView，这是一种 OPS 技术，它在原位测序之前利用体外转录来放大条形码，从而能够在包括原代细胞和组织在内的不同系统中进行高度多重表型读数的筛选。我们在动物模型的诱导多能干细胞衍生神经元、原代免疫细胞和肿瘤组织切片中展示了 PerturbView。在对原代骨髓源性巨噬细胞中的免疫信号通路进行筛选时，PerturbView 发现了 NF-κB 信号传导的已知和新型调节因子。此外，我们将 PerturbView 与小鼠异种移植模型组织切片中的空间转录组学相结合，为具有丰富光学和转录组表型的原位筛选铺平了道路。 PerturbView 将 OPS 的范围扩大到更广泛的模型和应用。© 2024。作者获得 Springer Nature America, Inc. 的独家许可。

Optical pooled screening (OPS) is a scalable method for linking image-based phenotypes with cellular perturbations. However, it has thus far been restricted to relatively low-plex phenotypic readouts in cancer cell lines in culture due to limitations associated with in situ sequencing of perturbation barcodes. Here, we develop PerturbView, an OPS technology that leverages in vitro transcription to amplify barcodes before in situ sequencing, enabling screens with highly multiplexed phenotypic readouts across diverse systems, including primary cells and tissues. We demonstrate PerturbView in induced pluripotent stem cell-derived neurons, primary immune cells and tumor tissue sections from animal models. In a screen of immune signaling pathways in primary bone marrow-derived macrophages, PerturbView uncovered both known and novel regulators of NF-κB signaling. Furthermore, we combine PerturbView with spatial transcriptomics in tissue sections from a mouse xenograft model, paving the way to in situ screens with rich optical and transcriptomic phenotypes. PerturbView broadens the scope of OPS to a wide range of models and applications.© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.