title: Tissue clearing for confocal imaging of native and bio-artificial skeletal muscle.
abstract: Novel clearing techniques have revolutionized three-dimensional confocal imaging of the brain without the need for physical tissue sectioning. We evaluated three clearing methods, ScaleA2, Clear(T2), and 3DISCO for visualizing native and tissue engineered muscle by confocal microscopy. We found that Clear(T2) treatment improved the depth of visualization of immunohistochemical staining slightly, but did not improve depth of visualization of endogenous green fluorescent protein (GFP). ScaleA2 preserved endogenous GFP signal better and permitted significantly deeper GFP imaging, but it was incompatible with tropomyosin immunohistochemical staining. 3DISCO treatment preserved both endogenous GFP and immunohistochemical staining, and permitted significantly deeper imaging. Clearing time for the 3DISCO procedure is short compared to ScaleA2 and Clear(T2). We suggest that 3DISCO is the preferable clearing method for native and tissue engineered skeletal muscle tissue.
authors.affiliation: Tissue Engineering Lab, Department of Development and Regeneration , KU Leuven campus Kulak, Kortrijk.
authors.forename: L
authors.initials: L
authors.lastname: Decroix
authors.forename: V
authors.initials: V
authors.lastname: Van Muylder
authors.forename: L
authors.initials: L
authors.lastname: Desender
authors.forename: M
authors.initials: M
authors.lastname: Sampaolesi
authors.forename: L
authors.initials: L
authors.lastname: Thorrez
mesh_terms: D000818:Animals; D021522:Bioartificial Organs; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D021621:Imaging, Three-Dimensional; D007150:Immunohistochemistry; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal / Q000379:methods*; D018482:Muscle, Skeletal / Q000033:anatomy & histology* / Q000378:metabolism*; D032448:Myoblasts, Skeletal / Q000166:cytology / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D023822:Tissue Engineering
chemical_list: D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins
keywords: bio-artificial; clearing; confocal microscopy; skeletal muscle; tissue engineering; visualization