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Optical clearing potential of immersion-based agents applied to thick mouse brain sections.

ABSTRACT: We have previously demonstrated that the use of a commercially-available immersion-based optical clearing agent (OCA) enables, within 3-6 hours, three-dimensional visualization of subsurface exogenous fluorescent and absorbing markers of vascular architecture and neurodegenerative disease in thick (0.5-1.0mm) mouse brain sections. Nonetheless, the relative performance of immersion-based OCAs has remained unknown. Here, we show that immersion of brain sections in specific OCAs (FocusClear, RIMS, sRIMS, or ScaleSQ) affects both their transparency and volume; the optical clearing effect occurs over the entire visible spectrum and is reversible; and that ScaleSQ had the highest optical clearing potential and increase in imaging depth of the four evaluated OCAs, albeit with the largest change in sample volume and a concomitant decrease in apparent microvascular density of the sample. These results suggest a rational, quantitative framework for screening and characterization of the impact of optical clearing, to streamline experimental design and enable a cost-benefit assessment.

SUBMITTER: Loren M

PROVIDER: S-EPMC6510422 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Optical clearing potential of immersion-based agents applied to thick mouse brain sections.

Loren Mathew M Crouzet Christian C Bahani Adrian A Vasilevko Vitaly V Choi Bernard B

PloS one 20190510 5

We have previously demonstrated that the use of a commercially-available immersion-based optical clearing agent (OCA) enables, within 3-6 hours, three-dimensional visualization of subsurface exogenous fluorescent and absorbing markers of vascular architecture and neurodegenerative disease in thick (0.5-1.0mm) mouse brain sections. Nonetheless, the relative performance of immersion-based OCAs has remained unknown. Here, we show that immersion of brain sections in specific OCAs (FocusClear, RIMS, ...[more]

PMID: 31075111

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Project description:BackgroundThe microvasculature is the network of blood vessels involved in delivering nutrients and gases necessary for tissue survival. Study of the microvasculature often involves immunohistological methods. While useful for visualizing microvasculature at the µm scale in specific regions of interest, immunohistology is not well suited to visualize the global microvascular architecture in an organ. Hence, use of immunohistology precludes visualization of the entire microvasculature of an organ, and thus impedes study of global changes in the microvasculature that occur in concert with changes in tissue due to various disease states. Therefore, there is a critical need for a simple, relatively rapid technique that will facilitate visualization of the microvascular network of an entire tissue.Methodology/principal findingsThe systemic vasculature of a mouse is stained with the fluorescent lipophilic dye DiI using a method called "vessel painting". The brain, or other organ of interest, is harvested and fixed in 4% paraformaldehyde. The organ is then sliced into 1 mm sections and optically cleared, or made transparent, using FocusClear, a proprietary optical clearing agent. After optical clearing, the DiI-labeled tissue microvasculature is imaged using confocal fluorescence microscopy and adjacent image stacks tiled together to produce a depth-encoded map of the microvasculature in the tissue slice. We demonstrated that the use of optical clearing enhances both the tissue imaging depth and the estimate of the vascular density. Using our "optical histology" technique, we visualized microvasculature in the mouse brain to a depth of 850 µm.Conclusions/significancePresented here are maps of the microvasculature in 1 mm thick slices of mouse brain. Using combined optical clearing and optical imaging techniques, we devised a methodology to enhance the visualization of the microvasculature in thick tissues. We believe this technique could potentially be used to generate a three-dimensional map of the microvasculature in an entire organ.

Dataset Information

Optical clearing potential of immersion-based agents applied to thick mouse brain sections.

Publications

Optical clearing potential of immersion-based agents applied to thick mouse brain sections.

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