title: ACT-PRESTO: Biological Tissue Clearing and Immunolabeling Methods for Volume Imaging.
abstract: The identification and exploration of the detailed organization of organs or of the whole body at the cellular level are fundamental challenges in biology. Transitional methods require a substantial amount of time and effort to obtain a 3D image and including sectioning the intact tissue, immunolabeling, and imaging serially-sectioned tissue, which produces a loss of information at each step of the process. In recently developed approaches for high-resolution imaging within intact tissue, molecular characterization has been restricted to the labeling of proteins. However, currently available protocols for organ clearing require a considerably long process time, making it difficult to implement tissue clearing techniques in the lab. We recently established a rapid and highly-reproducible protocol termed ACT-PRESTO (active clarity technique-pressure related efficient and stable transfer of macromolecules into organs), which allows for tissue clearance within several hours. Moreover, ACT-PRESTO enables rapid immunolabeling with conventional methods and accelerates antibody penetration into the deep layer of densely-formed, thick specimens by applying pressure or convection flow. We describe how to prepare tissues, how to clear by lipid removal using electrophoresis, and how to immuno-stain by a pressure-assisted delivery. The rapidity and consistency of the protocol will expedite the performance of 3D histological research and volume-based diagnoses.
authors.affiliation: Department of Anatomy, Korea University College of Medicine.
authors.forename: Eunsoo
authors.initials: E
authors.lastname: Lee
authors.affiliation: Department of Anatomy, Korea University College of Medicine; woongsun@korea.ac.kr.
authors.forename: Woong
authors.initials: W
authors.lastname: Sun
mesh_terms: D000818:Animals; D001921:Brain; D004586:Electrophoresis; D016591:Histocytological Preparation Techniques / Q000379:methods*; D020100:Hydrogels / Q000737:chemistry; D021621:Imaging, Three-Dimensional / Q000379:methods*; D007150:Immunohistochemistry / Q000379:methods*; D051379:Mice; D011312:Pressure
chemical_list: D020100:Hydrogels