Document Type
Article
Publication Date
10-10-2024
Abstract
We report a single-step optical clearing method that is compatible with RNA fluorescence in situ hybridization (FISH) imaging. We previously demonstrated microscopy imaging with immunohistochemistry and genetic reporters using a technique called lipid-preserving refractive index matching for prolonged imaging depth (LIMPID). Our protocol reliably produces high-resolution three-dimensional (3D) images with minimal aberrations using high magnification objectives, captures large field-of-view images of whole-mount tissues, and supports co-labeling with antibody and FISH probes. We also custom-designed FISH probes for quail embryos, demonstrating the ease of fabricating probes for use with less common animal models. Furthermore, we show high-quality 3D images using a conventional fluorescence microscope, without using more advanced depth sectioning instruments such as confocal or light-sheet microscopy. For broader adoption, we simplified and optimized 3D-LIMPID-FISH to minimize the barrier to entry, and we provide a detailed protocol to aid users with navigating the thick and thin of 3D microscopy.
Keywords
3D imaging, confocal microscopy, FISH, fluorescence in situ hybridization, light sheet microscopy, LIMPID, tissue clearing
Language
English
Publication Title
Journal of Biophotonics
Rights
© 2024 The Author(s). Journal of Biophotonics published by Wiley-VCH GmbH. This is an open access work distributed under the terms of the Creative Commons Attribution-Non-Commercial (https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License
Recommended Citation
Suh, J., Liu, Y., Smith, J., Watanabe, M., Rollins, A. and Jenkins, M. (2024), A Simple and Fast Optical Clearing Method for Whole-Mount Fluorescence In Situ Hybridization (FISH) Imaging. J. Biophotonics e202400258. https://doi.org/10.1002/jbio.202400258
Manuscript Version
Final Publisher Version