STEP 1. Tissue-Hydrogel Hybridization
A fixed tissue sample is incubated in the X-CLARITY™ Hydrogel Solution Kit to allow hydrogel monomers to diffuse uniformly throughout the sample. Once the solution has permeated the tissue, the sample is placed in the X-CLARITY™ Polymerization System. Monomers polymerize in the anaerobic environment, covalently linking the biomolecules to a sturdy hydrogel network, preserving molecular information and structural integrity.
STEP 2. Tissue Clearing
Once the tissue-hydrogel hybrid has been formed, the hybrid is cleared in the X-CLARITY™ Tissue Clearing System II using the ready-to-use Electrophoretic Tissue Clearing Solution. Lipids are extracted actively through electrophoresis or passively, leaving behind a stable and transparent tissue-hydrogel hybrid that is chemically accessible for molecular phenotyping.
STEP 3. Labeling
The transparent sample is then labled with antibodies using the DeepLabel Antibody Staining Kit, which enhances antibody penetration into clarified tissues. Optimized for clarified tissue samples, DeepLabel can be used with tissues cleared with various clearing methods such as passive or active CLARITY, iDISCO, 3DISCO, uDISCO, Visikol, or CUBIC.
STEP 4. Imaging
Prior to imaging, the tissue-hydrogel hybrid is placed in a refractive index (RI) matching solution in order to homogenize the environment within the tissue to the solution. This reduces light scatter, which in turn increases optical transparency and consequently increases image quality and imaging depth.