LifeCanvas Technologies is pioneering holistic intact-sample tissue processing with its sample-to-dataset pipeline and tools, seamlessly integrating each step in the workflow. As the only company to own the entire workflow, from tissue preservation, active clearing, antibody labeling, and refractive index-matching, to light-sheet imaging and image analysis, we are positioned to break down barriers and empower researchers with our streamlined approaches to ensure the best results and data possible.
Below you will find materials to help you make best use of LifeCanvas’s products, including the SHIELD tissue preservation protocol, tissue clearing resources, safety data sheets for reagents, and video tutorials.
Delipidating tissue to enable intact-sample labeling and imaging involves exposing samples to harsh treatments like high temperatures and pH changes. If left unchecked, this can cause damage to proteins and overall tissue structure, leading to incomplete or inaccurate image datasets. Therefore, to preserve the sample’s endogenous fluorescence and antigenicity along with tissue architecture, we use a novel tissue preservation technique called SHIELD that forms intramolecular bonds using polyfunctional, flexible epoxides to stabilize tissue architecture.
After SHIELD preservation, the tissue can be delipidated and either refractive index-matched for imaging or subjected to active immunohistochemical labeling. Notably, SHIELD-preserved tissues are well-suited for antibody multiplexing, i.e., iterative staining and de-staining, without loss of tissue antigenicity, to build up a rich picture of protein expression over repeated rounds of imaging. Check out the FAQ page for more information.
Full Pipeline Protocol (see “SHIELD” section beginning on page 3)
Eliminating membrane lipids is crucial to enabling better light penetration for imaging and increasing tissue permeability for active transport of molecular probes deep into intact tissue. Our active clearing device, SmartClear II Pro, employs a patent-pending stochastic electrotransport mechanism to foster rapid delivery of exogenous molecules such as the detergent SDS into tissues, facilitating uniform removal of light-scattering membrane lipids. Thanks to the application of a rotational electric field which minimizes the displacement of structural biomolecules, the tissue is cleared without any damage or deformation.
SmartClear II Pro is compatible with tissue samples preserved with SHIELD, CLARITY, and SWITCH. These methods avoid the loss of endogenous fluorescence and tissue shrinkage present in organic solvent based clearing methods like BABB, iDISCO, uDISCO, 3DISCO, and vDISCO. Compared to passive clearing methods, SmartClear II Pro offers orders of magnitude faster clearing with an easy-to-use interface to increase clearing throughput. We have continuously improved upon the original eletrophoretic clearing design, introducing stochastic electrotransport and a dual buffer-membrane system. These improvements ensure homogenous delipidation while preventing black particle formation all with economic and user-friendly buffers that last up to 10 days.
For first time users, a quick overview of how to get started using your device is below:
1. Preserve your tissue with a SHIELD tissue preservation kit. Perfusion is recommended, though you may also choose to perfuse with PFA and then post-fix the sample in SHIELD.
2. Install your SmartClear II Pro device, following the Quick Installation guide (below).
3. Read the User’s Manual (below) to familiarize yourself with the process. Feel free to contact us at firstname.lastname@example.org with any questions.
4. Start actively clearing your samples! Consult our support team and the FAQ page during your first clearing experiments to ensure you get the results you want as soon as possible. We can provide both technical and scientific support as well as feedback in judging tissue clarity so that you can get the data you need.
Full Pipeline Protocol (see “Clearing” section beginning on page 7)
After clearing and optional labeling, translucent, delipidated samples are incubated in EasyIndex, our refractive index (RI)-matching solution, raising and homogenizing their RI and turning them fully transparent. This increases light penetration into the sample and ensures that images will exhibit the highest resolution and focus. The cleared and labeled samples are then embedded in an agarose gel to stabilize them while imaging, ensuring alignment across channels and aiding colocalization of signals.
Full Pipeline protocol (see “Index Matching” section beginning on page 13)
Save space, time, and money with EasyClear. The intuitive touch-screen interface allows the user to precisely control temperature (20-90°C), shaking speed (0-60 rpm), and tilting angle (0-22°) for optimal tissue processing of up to 10 samples simultaneously. Leave behind the evaporation and contamination issues common in water baths, incubators, and shakers. EasyClear is ideal for tissue clearing, tissue staining, or any buffer washing.
EasyGel provides complete and consistent vacuuming/N2 purging of up to 8 samples simultaneously with vacuum tubes that connect to each sample tube. The system also allows the user to precisely control temperature and shaking speed.
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