Cryosectioning fresh frozen tissues for multimodal imaging

Clean the cryostat surface, anti-roll blade and forceps with ethanol before sectioning.

Place the tissue block inside the cryostat at -20°C for 30 mins for equilibration. Also, place the slides and substrates inside the cryostat for cooling.

Set the chamber temperature of the cryostat to -20C and object temperature to -18C to -20C

Place the blade in place and lock it.

Trim the tissue block at 30-50µm thickness to obtain a flat surface.

Mount the tissue block on the chuck using water (no OCT) and adjust the angle of the chuck as needed.

Position the anti-roll bar appropriately so that it doesn’t scrape the tissue (if placed too far) or roll off of the block (if placed too near).

Gently section the tissue at 8µm thickness using an anti-roll bar and place the consecutive sections on individual substrates. Adjust and flatten with a fine tipped brush if necessary. For H&E, DESI and RNAscope imaging, tissue sections are placed on regular or positively charged microscope slides and on gold coated 2.5cm² substrate for SIMS-imaging.

Thaw mount the sections on the target using heat from the finger under the slide.

Store at -80C until use.