Question: How should I prepare my FFPE block upstream of the Isolation of Cells from FFPE Tissue Sections for Chromium Fixed RNA Profiling Demonstrated Protocol?
Answer: We would recommend following best practices outlined in our Visium Spatial Gene Expression for FFPE assay documentation. Below are general recommendations:
Block generation: If you have access to fresh tissue or have the ability to control processing methods, we would advise using fresh or frozen tissue and performing either the Chop/Fix Demonstrated Protocol or utilizing the Chromium Nuclei Isolation Kit instead of generating FFPE blocks. Fresh or frozen tissue that’s subsequently PFA fixed will maintain the RNA quality better than FFPE.
Block storage: FFPE blocks are typically stored at room temperature but storage at 4°C and protected from light is ideal when using tissue blocks for NGS analysis, as this will slow the degradation of RNA.
Sectioning blocks: Practice sectioning prior to generating curls for your Fixed RNA Profiling experiment. Before sectioning, clean the microtome and water bath with RNase Decontamination Solution. Be sure to utilize a new, clean blade (free of oil) to ensure high-quality curls are generated. When transferring FFPE curls to tubes for deparaffinization and rehydration, transfer them gently so as to not damage or break them. During the Isolation of Cells from FFPE Tissue Sections for Chromium Fixed RNA Profiling Demonstrated Protocol steps, use validated nuclease-free reagents, consumables, and tools.
See the following documents for more guidance on isolation and FFPE block sectioning with our Visium products:
- Isolation of Cells from FFPE Tissue Sections for Chromium Fixed RNA Profiling Demonstrated Protocol
- Visium Spatial Gene Expression for FFPE Tissue Preparation Guides
Products: Fixed RNA Profiling