Loading specimens into SLICE

Overview

Cleared-tissue specimens must be immobilized inside a SLICE Cuvette, containing mounting medium and sealed with a SLICE Cuvette quick-release mount in order to image them on a SLICE system. Position and immobilize the sample inside the cuvette using a Brain Bracket, then submerge in storage solution (that is compatible with the clearing method used to prepare the sample) to maintain clarity and prevent drift during imaging. It is not strictly required to use a Brain Bracket, however, they enable convenient tissue handling and stable positioning. A thin layer of silicone sealant applied at the cuvette‑mount junction can prevent mounting medium from evaporating and is recommended for imaging and storing samples. Once assembled, secure the sealed cuvette onto the SLICE microscope stage; SLICE Cuvette quick-release mounts are stably held on the stage by magnets.

Required materials

  • SLICE cuvettte, any size, MBF Bioscience, containing the following:

    • Fused quartz glass cuvette (there are several sizes)

    • SLICE quick-release coupling mount (this is the lid for the cuvette that both seals the cuvette and magnetically connects the cuvette to the SLICE stage.

    • Brain bracket (available soon)

  • Cleared-tissue specimen, sized to fit in the cuvette. Compatible clearing methods include iDISCO, CLARITY, SHANEL, CUBIC, BINAREE and other optical clearing techniques.

  • Storage solution appropriate for the tissue-clearing method

  • Clear silicone waterproof sealant. We have used this sealant, but any similar product will work.

  • (optional) Thin, heat resistant, transparent, silicone sheet, cut into slices that fit into the cuvette you will be using. We have used this product.

Chemical risks to people and equipment

Many chemicals used in tissue-clearing protocols are hazardous and potentially volatile. To minimize risk, we highly recommend sealing the cuvette with silicone sealant after mounting samples.

Note that some of these chemicals may also be corrosive, presenting a hazard to SLICE hardware. Avoid spilling or dripping corrosive material on the instrument; clean up immediately if spills or drips do occur.

Procedure

Employ chemical-safety precautions, such as wearing personal protective equipment appropriate for the chemicals present in your cleared specimen.

A. Use a Brain bracket to place sample in the cuvette

  1. Mount cleared tissue on an MBF Bioscience Brain bracket.

    1. Gently remove storage solution from cleared tissue using a lint-free tissue.

    2. Apply small amount of silicone sealant to the bottom and back surfaces of the Brain Bracket.

    3. Position the cleared tissue onto the sealant in the correct orientation.

    4. Apply gentle pressure for 5 minutes to secure the tissue.

    5. Allow to dry for 1 hour until cured.

  2. Insert Brain Bracket into cuvette

    1. Gently insert the Brain Bracket into the cuvette and settle it fully at the bottom.

    2. Fill the cuvette with mounting medium until brain tissue is fully immersed.

    3. Pipette out any bubbles in the medium.

alt A. Place sample in the cuvette without using a brain bracket

  1. Place the brain into the cuvette

    1. Add a small amount of storage solution to the bottom of the cuvette.

    2. Carefully place the brain sample into the cuvette and adjust its orientation for imaging. Minimize the time brain is exposed to air so that it does not become dried out.

    3. Add more storage solution to fully immerse the brain tissue.

  2. Add pieces of silicone sheet to immobilize the tissue.

    1. Gently push the brain toward the side of the cuvette that will face the detection objective.

    2. Stack several silicone slices to form a block that will fill the space between the brain and the cuvette surface opposite the detection objective.

      Trim the silicone slices so that the block fits into the cuvette without scratching it.

    3. Insert the trimmed silicone block into the cuvette to hold the brain in position.

    4. Remove any bubbles that may have formed in the cuvette and wipe away any storage solution on the exterior of the cuvette.

B. Seal the cuvette and attach it to the SLICE stage

  1. Seal the cuvette with a SLICE quick-release coupling mount.

    1. Place the cuvette mount onto the cuvette with gentle pressure. Avoid using excessive force to prevent breaking or cracking the cuvette.

    2. Apply a thin layer of silicone sealant around the junction to secure the mount.

    3. Allow to dry for 1 hour until cured.

  2. Attach cuvette to the SLICE stage.

    1. Tilt the cuvette slightly and hook the right slot of the magnet mount onto the right stage rod from below.

    2. Use the right rod as a pivot and rotate the cuvette clockwise until the left slot engages with the left rod.