Generalizable virtual staining from histopathology to spatial proteomics across biological scales

Abstract

Large histology archives capture morphological correlates of molecular tissue states but lack direct spatial protein measurements. In this study, we show that routine hematoxylin and eosin (H&E) histology can provide a queryable, morphology-based representation of spatial protein expression that remains informative from pixels to patients. To achieve this, we developed QUEST, a model framework that represents protein markers as semantic queries for flexible multiplex immunofluorescence (MIF) prediction. Trained on a heterogeneous atlas of paired H&E and MIF images comprising over 10,000 tissue regions, QUEST captured morphological patterns associated with protein markers and reconstructed their expression intensities across diverse tissue types and cohorts. Its utility extended beyond image reconstruction and preserved biologically meaningful information across multiple scales: virtual staining preserved protein marker expression at the cellular and tissue scales, supported cell phenotyping, cellular neighborhood analysis, and characterization of spatial tissue organization, retained clinically relevant signals for patient stratification, and enabled H&E-based queries to retrieve MIF references with similar molecular profiles. These findings position virtual staining not simply as image generation but as a flexible interface between routine histology and spatial proteomics, expanding the analytical value of existing H&E archives.