LifeCanvas SmartLabel rapid staining of mouse brain hemi with ThermoFisher anti-GFP (A-11122).

< 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.

LifeCanvas SmartLabel rapid immunostaining of mouse brain hemi with anti-Parvalbumin. 

< 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.

GFAP immunostaining in a 2-mm tissue slab reveals astrocytes’ lining of vasculature.

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).

tdTomato labeling of a mouse spinal cord cell-type.

Cleared with SmartClear II Pro and imaged with SmartSPIM.

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

Intravenously-delivered fluorescent dye traces out mouse brain blood vessels.

Mouse brain labeled with vascularly delivered fluorescent dye.

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

Image courtesy of Kleinfeld and Lim labs, UCSD.

Fluorescent nuclear dye staining of an entire mouse brain hemisphere in < 2 days.

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.

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).

Calb2:tdTomato in mouse brain.

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.

Visualizing cerebellar circuits with tdTomato.

Cleared with SmartClear II Pro and imaged with SmartSPIM.

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