Optimization of GFP Fluorescence Preservation by a Modified uDISCO Clearing Protocol

Frontiers in Neuroanatomy
Yusha LiDan Zhu

Abstract

Tissue optical clearing techniques provide alternative approaches for imaging large-volume specimens. uDISCO, an organic-solvent-based method, stands out from the enormous array of available optical clearing methods by achieving whole-brain imaging with high transparency, size reduction and fluorescence preservation. In this study, we aimed to modify the uDISCO protocol to achieve better fluorescence preservation and to thereby further improve its optical imaging quality. First, we determined the optimal pH value for optimized uDISCO, termed "a-uDISCO" (alkaline pH-based uDISCO). Then, we compared fluorescence preservation between a-uDISCO and uDISCO. In addition, we validated the clearing performance of the optimized method according to several parameters, including tissue transparency, size changes, and the maintenance of cell morphology. Finally, we demonstrated that a-uDISCO enabled the high-quality brain-wide visualization of neuronal structures. This method potentially provides a better alternative for high-throughput imaging of samples with low-level fluorescence protein expression or for archiving and repetitive revisiting of rare samples.

References

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Citations

Feb 13, 2019·Science Advances·Yisong QiDan Zhu
Jan 19, 2020·The Journal of Immunology : Official Journal of the American Association of Immunologists·Paweł MatrybaJakub Gołąb
May 15, 2020·Chemical Communications : Chem Comm·Quan XuMeng Xu
Sep 18, 2020·Journal of Anatomy·Ting TianXiaoguang Li
Nov 2, 2019·Scientific Reports·Deshka S FosterMichael T Longaker
Mar 27, 2021·Molecular Systems Biology·Muge MolbayAli Ertürk
Mar 15, 2021·Journal of Biophotonics·Yanjing ZhanShiwen Zhang
Jul 3, 2021·International Journal of Molecular Sciences·Paweł MatrybaJakub Gołąb
Aug 7, 2021·Progress in Biophysics and Molecular Biology·Camilla OliantiLeonardo Sacconi
Dec 4, 2021·ELife·Benoît BoulanJean-Christophe Deloulme

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Methods Mentioned

BETA
transgenic
Fluorescence
Light-sheet

Key Resources (RRID) Mentioned

IMSR_JAX
SCR_003070
SCR_007370
SCR_002865

Software Mentioned

OPTIClear
SPSS
uDISCO
Imaris
ImageJ
CUBIC
CLARITY

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