Single-cell analyses have revealed extensive heterogeneity between and within human tumours^1,2,3,4, but complex single-cell phenotypes and their spatial context are not at present reflected in the histological stratification that is the foundation of many clinical decisions. Here we use imaging mass cytometry⁵ to simultaneously quantify 35 biomarkers, resulting in 720 high-dimensional pathology images of tumour tissue from 352 patients with breast cancer, with long-term survival data available for 281 patients. Spatially resolved, single-cell analysis identified the phenotypes of tumour and stromal single cells, their organization and their heterogeneity, and enabled the cellular architecture of breast cancer tissue to be characterized on the basis of cellular composition and tissue organization. Our analysis reveals multicellular features of the tumour microenvironment and novel subgroups of breast cancer that are associated with distinct clinical outcomes. Thus, spatially resolved, single-cell analysis can characterize intratumour phenotypic heterogeneity in a disease-relevant manner, with the potential to inform patient-specific diagnosis.

单细胞分析揭示了人类肿瘤之间和内部的广泛异质性^1,2,3,4 ，但复杂的单细胞表型及其空间背景目前尚未反映在作为许多临床决策基础的组织学分层中。在这里，我们使用成像质谱流式细胞术⁵同时量化 35 种生物标志物，获得 352 名乳腺癌患者的肿瘤组织的 720 张高维病理图像，以及 281 名患者的长期生存数据。空间分辨的单细胞分析确定了肿瘤和基质单细胞的表型、它们的组织和异质性，并使得能够根据细胞组成和组织组织来表征乳腺癌组织的细胞结构。我们的分析揭示了肿瘤微环境的多细胞特征以及与不同临床结果相关的乳腺癌新亚组。因此，空间分辨的单细胞分析可以以疾病相关的方式表征肿瘤内表型异质性，并有可能为患者特异性诊断提供信息。