Protocol for ultrafast immunolabeling and 3D imaging of whole organs and large tissues

High-resolution whole-organ imaging provides cellular and molecular insights into tissue and tumor microenvironments. Here, we present a protocol for processing whole soft tissue organs and human tissues for 3D imaging within 2–2.5 days. We describe steps for bleaching, antigen retrieval, and collagenase-based digestion, followed by whole-tissue immunolabeling. We then detail procedures for dehydration and optical clearing for imaging by light-sheet microscopy. For complete details on the use and execution of this protocol, please refer to Biswas et al. 1