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Updated: Jun 7, 2026
ACT-PRESTO: Biological Tissue Clearing and Immunolabeling Methods for Volume Imaging
Published on: December 31, 2016
Protocol for ultrafast immunolabeling and 3D imaging of whole organs and large tissues
Zhangfan Ding1, Hanyu Liu2, Junyu Chen3
1Tissue and Tumor Microenvironments Lab, Cancer Discovery and Regenerative Medicine Program, Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore 636921, Singapore; State Key Laboratory of Oral Diseases, National Center for Stomatology, National Clinical Research Center for Oral Diseases, Department of Head and Neck Oncology, West China Hospital of Stomatology, Sichuan University, Chengdu 610041, China.
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.
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.
