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Effects of Clearing Media on Tissue Transparency and Shrinkage

Light sheet imaging of insect brains cleared with media having a tunable refractive index

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This study comprehensively evaluates the effects of different clearing media on tissue transparency and shrinkage by comparing freshly dissected dipteran fly brains with their cleared equivalents. Tissue clearing combined with light sheet microscopy has become a powerful 3D imaging and quantification tool for whole organs. As tissue clearing facilitates subcellular-level optical access to intact tissue, it has the potential to visualize previously unseen details of complex organs like the brain. For high-resolution imaging of thick samples, the clearing medium needs to match the high refractive index of the immersion liquids used with high-NA objectives. This point is of particular importance in insect brains, where numerous (initially air-filled) tracheal tubes are branching throughout the brain and exacerbate any scattering of light. 

Brains were imaged with the Digital LightSheet (DLS) from Leica Microsystems. It allows the user to choose between several detection objectives and mounting frames enabling high-resolution visualization. Users can also take full advantage of all the structural details revealed within the 3D cleared insect brains.

 

Figure 1:Illustration showing sample mounting and imaging on the DLS.
Figure 2: Comparison of Leica SP8 confocal (20x oil immersion objective) (top) and Leica SP8 DLS imaging (5x/0.15 objective and DLS TwinFlect 7.8 mm mirrors) (bottom) of Rapiclear1.49E cleared whole insect brain with Lucifer yellow injected wide-field motion neuron.

Read the full article:

Bekkouche B.M.B., Fritz H.K.M., Rigosi E. & O'Carroll D.C.:

Comparison of Transparency and Shrinkage During Clearing of Insect Brains Using Media With Tunable Refractive Index

Front. Neuroanat., 20 November 2020, https://doi.org/10.3389/fnana.2020.599282

https://www.frontiersin.org/articles/10.3389/fnana.2020.599282/full

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