Abstract
Spatially resolved transcriptomics (SRT) studies are becoming increasingly common and large, offering unprecedented opportunities in mapping complex tissue structures and functions. Here we present integrative and reference-informed tissue segmentation (IRIS), a computational method designed to characterize tissue spatial organization in SRT studies through accurately and efficiently detecting spatial domains. IRIS uniquely leverages single-cell RNA sequencing data for reference-informed detection of biologically interpretable spatial domains, integrating multiple SRT slices while explicitly considering correlations both within and across slices. We demonstrate the advantages of IRIS through in-depth analysis of six SRT datasets encompassing diverse technologies, tissues, species and resolutions. In these applications, IRIS achieves substantial accuracy gains (39-1,083%) and speed improvements (4.6-666.0) in moderate-sized datasets, while representing the only method applicable for large datasets including Stereo-seq and 10x Xenium. As a result, IRIS reveals intricate brain structures, uncovers tumor microenvironment heterogeneity and detects structural changes in diabetes-affected testis, all with exceptional speed and accuracy.