Fig. 1: Exemplar framework for good practice in tissue preparation and sectioning.

A stepwise approach for tissue preparation and sectioning for spatial transcriptomic (ST) analysis (FFPE sections of BMT tissue for 10x Xenium analysis). Prior to sectioning the environment should be cleaned with RNaseZap to ensure a RNase free environment. 1. Pathology review of a H&E-stained section. 2. Review FFPE tissue blocks to ensure macroscopic BMT tissue integrity. 3. Gently and very lightly score around the BMT region of interest with a scalpel, excluding areas not suitable for ST analysis e.g. thick cortical bone/haemorrhage. 4. Chill and rehydrate the FFPE block in an ice bath and then cut and discard the first few sections until a full-face of the BMT is seen and surface paraffin is removed. Cut a thin (see relevant protocol for thickness) section from the FFPE block with a microtome. 5. Immediately transfer the tissue section to surface of a warm (42°C or as per protocol) water bath using tweezers, careful to touch only the peripheral paraffin rather than tissue. Lie the section flat on the surface of the water and inspect the section for integrity and absence of folding. 6. Without touching the BMT tissue, remove the excess paraffin from the section with tweezers and discard. The light scoring in step three will facilitate removal of excess paraffin whilst preserving the BMT tissue. 7. Manoeuvre the BMT sections (on the surface of the water) onto the appropriate region of the ST slide. Placing the ST slide in the water bath under the tissue sections minimises the need to touch the tissue during this step. 8. Allow the tissue to dry on the slide as per protocol to be taken forward for further processing. Refer to the relevant user manual for further details including regarding water bath preparation, block chilling, drying temperature and timing.