Tutorial: methods for three-dimensional visualization of archival tissue material

Analysis of three-dimensional patient specimens is gaining increasing relevance for understanding the principles of tissue structure as well as the biology and mechanisms underlying disease. New technologies are improving our ability to visualize large volume of tissues with subcellular resolution. One resource often overlooked is archival tissue maintained for decades in hospitals and research archives around the world. Accessing the wealth of information stored within these samples requires the use of appropriate methods. This tutorial introduces the range of sample preparation and microscopy approaches available for three-dimensional visualization of archival tissue. We summarize key aspects of the relevant techniques and common issues encountered when using archival tissue, including registration and antibody penetration. We also discuss analysis pipelines required to process, visualize and analyze the data and criteria to guide decision-making. The methods outlined in this tutorial provide an important and sustainable avenue for validating three-dimensional tissue organization and mechanisms of disease.