Antigen retrieval and clearing for whole-organ immunofluorescence by FLASH.
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Embargo End Date
ICR Authors
Authors
Messal, HA
Almagro, J
Zaw Thin, M
Tedeschi, A
Ciccarelli, A
Blackie, L
Anderson, KI
Miguel-Aliaga, I
van Rheenen, J
Behrens, A
Almagro, J
Zaw Thin, M
Tedeschi, A
Ciccarelli, A
Blackie, L
Anderson, KI
Miguel-Aliaga, I
van Rheenen, J
Behrens, A
Document Type
Journal Article
Date
2020-11-27
Date Accepted
2020-09-18
Abstract
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.
Citation
Nature Protocols, 2020, 16 (1), pp. 239 - 262
Source Title
Nature Protocols
Publisher
NATURE PORTFOLIO
ISSN
1754-2189
eISSN
1750-2799
1750-2799
1750-2799
Collections
Research Team
Cancer Stem Cell
Convergence SC Management
Convergence SC Management