An end-to-end workflow for non-destructive 3D pathology.

• Type of Material: Article
• Authors: Kevin W. Bishop, University of Washington; Lindsey A. Erion Barner, University of Washington; Qinghua Han, University of Washington; Elena Baraznenok, University of Washington; Lydia Lan, University of Washington; Chetan Poudel, University of Washington; Gan Gao, University of Washington; Robert B. Serafin, University of Washington; Sarah S. L. Chow, University of Washington; Adam K. Glaser, University of Washington; Andrew Janowczyk, Emory University; David Brenes, University of Washington; Hongyi Huang, University of Washington; Dominie Miyasato, University of Washington; Lawrence D. True, University of Washington; Soyoung Kang, University of Washington; Joshua C. Vaughan, University of Washington; Jonathan T. C. Liu, University of Washington
• Language: English
• Date: 2023-08-06
• Publisher: NIH
• Publication Version: Preprint (Prior to Peer Review)
• Copyright Statement: The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.
• License: Creative Commons Attribution-NonCommercial 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1101/2023.08.03.551845
• Title of Journal or Parent Work: bioRxiv
• Grant/Funding Information: This work was supported by funding from the National Institutes of Health (NIH): R01EB031002 and R01CA268207 (J.T.C.L.); R00CA240681 (A.K.G.); and U01CA239055, 1R01LM013864, 1U01DK133090, U01CA248226, and 2R01DK118431 (A.J.); the National Science Foundation (NSF): 1934292 HDR: I-DIRSE-FW (J.T.C.L.) and NSF Graduate Research Fellowships DGE-1762114 (K.W.B. and L.A.E.B.); the Department of Defense (DoD) Prostate Cancer Research Program: W81XWH-20-1-0851 (J.T.C.L.) and W81XWH-18-10358 (J.T.C.L. and L.D.T); Prostate Cancer UK: MA-ETNA19-005 (J.T.C.L.), and the Pacific Northwest Prostate Cancer SPORE (P50CA097186).
• Keywords: 3D pathology; protocol; tissue; imaging; volumetric context
• Research Categories: Health Sciences, Radiology; Health Sciences, Pathology

Abstract

Recent advances in 3D pathology offer the ability to image orders-of-magnitude more tissue than conventional pathology while providing a volumetric context that is lacking with 2D tissue sections, all without requiring destructive tissue sectioning. Generating high-quality 3D pathology datasets on a consistent basis is non-trivial, requiring careful attention to many details regarding tissue preparation, imaging, and data/image processing in an iterative process. Here we provide an end-to-end protocol covering all aspects of a 3D pathology workflow (using light-sheet microscopy as an illustrative imaging platform) with sufficient detail to perform well-controlled preclinical and clinical studies. While 3D pathology is compatible with diverse staining protocols and computationally generated color palettes for visual analysis, this protocol will focus on a fluorescent analog of hematoxylin and eosin (H&E), which remains the most common stain for gold-standard diagnostic determinations. We present our guidelines for a broad range of end-users (e.g., biologists, clinical researchers, and engineers) in a simple tutorial format.