Cleared with SmartClear II Pro and imaged with SmartSPIM.

Sample courtesy of G. Allan Johnson, Duke Center for In Vivo Microscopy.

Whole mouse brain processed with LifeCanvas’ tissue processing pipeline. Preserved with SHIELD, cleared with SmartClear II Pro, labeled with SmartLabel with anti-ChAT, and imaged with SmartSPIM at 4 µm z-step and 1.8 µm pixel size. Tissue courtesy of the Fishell Lab at Broad Institute.

Mouse brain hemisphere displaying ChAT-positive neurons in both the brainstem and basal forebrain and dense labeling of Neuropeptide Y throughout the hypothalamus. Tissue was SHIELD perfused, cleared with SmartClear II Pro, and immunolabeled with SmartLabel with anti-NPY (cyan) and anti-ChAT (magenta). Imaged using SmartSPIM at 4 µm Z-step and 1.8 µm XY pixel size. Tissue courtesy of Hongwei Dong, Bin Zhang, and Neda Khanjani and the Center for Integrative Connectomics (CIC) lab.

Mouse duodenum segment, perfused with lectin (red) to label vasculature, SHIELD preserved, cleared with SmartClear II Pro, and immunolabeled with SmartLabel (anti-Olfm4 stem cell marker, cyan). Imaged in one piece using SmartSPIM at 2 µm Z-step and 1.8 µm XY pixel size.

Nuclear stain (SYTO16, in cyan) and vasculature stain (DyLight 649-conjugated tomato lectin, in pink) made possible in <1 day by SmartLabel. Tissue provided by: Translational Bioimaging Group @ Barrow Neurological Institute.

< 2-day immunolabeling of an intact mouse brain hemisphere using SmartLabel from LifeCanvas Technologies and ThermoFisher anti-GFP antibody (A-11122)! Tissue was perfused with LifeCanvas’s SHIELD (Park et al., Nature Biotech, 2018) solution kit, enabling robust preservation of endogenous fluorescent proteins, antigenicity, and overall tissue architecture. Tissue was next delipidated in SmartClear II Pro and then labeled actively with just 3 μl anti-GFP primary (A-11122) & a Fab fragment secondary (2.3 μl goat anti-rabbit Rhodamine Red™-X AffiniPure Fc-fragment specific, from Jackson ImmunoResearch) in SmartLabel, which uses a combination of stochastic electrotransport (Kim et al., PNAS, 2015) and the SWITCH method (Murray et al., Cell, 2015) of controlling molecular labeling reactions to immunostain intact samples in <2 days. Tissue was then refractive index matched using EasyIndex and imaged at single-cell resolution (1.8 μm/pixel in XY with 4-μm Z-step) on LifeCanvas’s SmartSPIM light-sheet microscope in only 60 minutes.

< 2-day immunolabeling of an intact mouse brain hemisphere using SmartLabel from LifeCanvas Technologies and ThermoFisher anti-Parvalbumin (PV) antibody (PA1-933)! Tissue was perfused with LifeCanvas’s SHIELD (Park et al., Nature Biotech, 2018) solution kit, enabling robust preservation of antigenicity and overall tissue architecture. Tissue was next delipidated in SmartClear II Pro and then labeled actively with 20 µg anti-PV primary & a Fab fragment secondary in SmartLabel, which uses a combination of stochastic electrotransport (Kim et al., PNAS, 2015) and the SWITCH method (Murray et al., Cell, 2015) of controlling molecular labeling reactions to immunostain intact samples in < 2 days. Tissue was then refractive index matched using EasyIndex and imaged at single-cell resolution (1.8 µm/pixel in XY with 4 µm Z-step; 561 nm laser line) on LifeCanvas’s SmartSPIM light-sheet microscope in only ~30 minutes.

2-mm thick slab of mouse brain immunostained for GFAP in < 2 days using SmartLabel.

Cleared with SmartClear II Pro, labeled with SmartLabel, and imaged with SmartSPIM (< 5 mins, 1.8 µm/pixel in XY with 2 µm Z-steps).

Cleared with SmartClear II Pro and imaged with SmartSPIM.

Sample courtesy of Prof. Helen Lai’s lab (helenlailab.org), UT Southwestern.

Mouse brain labeled with vascularly delivered fluorescent dye.

Cleared with SmartClear and imaged with SmartSPIM (10x).

Image courtesy of Kleinfeld and Lim labs, UCSD.

Using LifeCanvas’s SmartLabel platform, green fluorescent nuclear dye (SYTO 16) staining of a mouse cerebral hemisphere is possible in < 2 days, with labeling density & intensity that is uniform from the tissue’s surface to its core. Along with labeling of other cell-types, staining of astrocytes traces out the brain’s intricate network of vasculature.

Cleared with SmartClear II Pro, labeled with SmartLabel (< 2 days), and imaged with SmartSPIM (1.8 µm/pixel in XY with 4 µm Z-step, ~30 mins).

Thy1-driven YFP produces a fluorescent Golgi stain in an optically-clear mouse brain hemisphere, highlighting major brain pathways.

Cleared with SmartClear II Pro and imaged with SmartSPIM (~30 mins, 1.8 µm/pixel in XY with 2 µm Z-steps).

tdTomato expression driven by the Calb2 promoter reveals circuits formed by cerebellar Purkinje neurons, sub-types of inhibitory interneurons, and other cell-types in a mouse brain hemisphere.

Cleared with SmartClear II Pro and imaged with SmartSPIM (~30 minutes, 1.8 µm/pixel in XY with 4 µm Z-step).

Sample courtesy of Prof. Wei Xu’s lab, UT Southwestern.

Cleared with SmartClear II Pro and imaged with SmartSPIM.

Sample courtesy of Prof. Helen Lai’s lab (helenlailab.org), UT Southwestern.