Logos Biosystems X-CLARITY for preserving protein signals for volumetric imaging of intact tissue

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As scientists continue to study disease states in a variety of tissues, an understanding of the 3D environment of organs is providing insight into structure–function relationships that affect tissue processes. Tissue clearing techniques of complete organs have replaced the need to reduce samples to thin serial sections histologically, which are giving a better view into the inner workings of complex tissues; however, there are still significant obstacles that stand in the way. Major limitations, particularly for solvent-based techniques, include degradation of the proteins sought to image, preserving the signal from endogenous fluorescent proteins (FPs), and limited solvent compatible probes.

The X-CLARITY from Logos Biosystems has been developed to standardise, simplify and accelerate the tissue clearing process. It is an easy-to-use, aqueous-based method that accelerates the removal of lipids from tissues in a simple, rapid and efficient manner — a whole rat brain can be cleared in just 6 h. Reproducible clearing results aid subsequent 3D and fluorescence imaging — particularly for confocal, light sheet and two-photon, which demand greater clearing depth. Said to be unmatched by other techniques, the X-CLARITY is an all-in-one system with ready-to-use reagents.

X-CLARITY makes clearing a turnkey operation of four steps: polymerise, clear, label and Refractive Index Match (RIM Media). Preserved tissues are embedded with a hydrogel matrix; lipids are actively extracted with electrophoresis to create a stable and optically transparent tissue-hydrogel hybrid that is chemically accessible for multiple rounds of antibody labelling and imaging. Once labelled, the tissue is placed in a RIMM to further enhance the clarity for microscopy. Native cytoarchitecture remains intact and even endogenous fluorescence proteins are preserved for robust fluorescence imaging downstream. Applications for the X-CLARITY method extend beyond brains to clear virtually any organ, even organoids.

The X-CLARITY system has three main components. A polymeriser activates the temperature-triggered hydrogel, which supports the structure of the tissue after fat removal maintaining shape and size. The electrophoretic tissue clearing chamber uses two large platinum electrode plates that maximise surface area and enable even clearing while pumping an aqueous detergent through the chamber. The control tower contains pumps and temperature regulation essential to protect proteins.

For more information: https://www.atascientific.com.au/products/logos-biosystems-x-clarity/.