Integrated light and electron microscopy workflow for morphological, molecular and ultrastructural analysis of spheroids

The dataset presented here supports an integrated light and electron microscopy workflow for comprehensive morphological, molecular, and ultrastructural characterization of three-dimensional (3D) spheroid models. The data accompany a submitted data paper and are discussed in a submitted original research article, with the aim of enabling standardized and reproducible multi-scale imaging of spheroids in research in vitro. The associated research article presents optimized workflows for light and electron microscopy analysis of spheroids derived from the human noncancerous urothelial cell line SV-HUC-1 and the malignant urothelial cancer cell line T24. The study describes protocols for histological staining and immunofluorescence on cryosectioned and paraffin-embedded spheroids, as well as whole-mount immunofluorescence combined with optical clearing and confocal microscopy for three-dimensional protein localization. Scanning and transmission electron microscopy further provide high-resolution insights into spheroid surface morphology and internal ultrastructure. Together, these complementary approaches establish a standardized framework for multi-scale spheroid characterization applicable across diverse in vitro models.