Antigen retrieval and clearing for whole-organ immunofluorescence by FLASH.
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Date
2020-11-27Author
Messal, HA
Almagro, J
Zaw Thin, M
Tedeschi, A
Ciccarelli, A
Blackie, L
Anderson, KI
Miguel-Aliaga, I
van Rheenen, J
Behrens, A
Type
Journal Article
Metadata
Show full item recordAbstract
Advances in light-sheet and confocal microscopy now allow imaging of cleared large biological tissue samples and enable the 3D appreciation of cell and protein localization in their native organ environment. However, the sample preparations for such imaging are often onerous, and their capability for antigen detection is limited. Here, we describe FLASH (fast light-microscopic analysis of antibody-stained whole organs), a simple, rapid, fully customizable technique for molecular phenotyping of intact tissue volumes. FLASH utilizes non-degradative epitope recovery and membrane solubilization to enable the detection of a multitude of membranous, cytoplasmic and nuclear antigens in whole mouse organs and embryos, human biopsies, organoids and Drosophila. Retrieval and immunolabeling of epithelial markers, an obstacle for previous clearing techniques, can be achieved with FLASH. Upon volumetric imaging, FLASH-processed samples preserve their architecture and integrity and can be paraffin-embedded for subsequent histopathological analysis. The technique can be performed by scientists trained in light microscopy and yields results in <1 week.
Collections
Subject
Animals
Antigens
Drosophila
Epitopes
Female
Fluorescent Antibody Technique
Humans
Imaging, Three-Dimensional
Kidney
Lacrimal Apparatus
Liver
Lung
Male
Mammary Glands, Human
Mice
Microscopy, Confocal
Organoids
Pancreas
Stomach
Research team
Cancer Stem Cell
Convergence SC Management
Language
eng
Date accepted
2020-09-18
License start date
2020-11-27
Citation
Nature Protocols, 2020, 16 (1), pp. 239 - 262
Publisher
NATURE RESEARCH