Question: Why are some tissue types or blocks more prone to lower usable reads in a Visium Spatial assay?
Answer: Tissue block composition, isolation method, and fixation quality can all greatly impact the quality of a Visium experiment.
For instance, if the tissue of interest is primarily fat or connective tissue, we expect that those regions of the tissue will have poor Gene Expression and Protein Expression signal (if using the Human Immune Cell Profiling Panel). Another consideration is how tissue is surgically isolated, in that if tissue is isolated via surgical resection rather than excision, there’s a higher likelihood of isolating excess and unwanted tissue along with the region of interest. As a consequence, this can lead to a larger proportion of reads coming from unwanted regions of the tissue and higher wasted reads. An additional consideration is how quickly and optimally the tissue is fixed. Fixation delays can negatively impact nucleic acid quality and preservation of tissue morphology, while over-fixation can lead to inefficient antigen retrieval and thus, lower sensitivity Gene and Protein Expression data.
Internally, we’ve found that spleen and kidney can experience lower usable Gene Expression reads due to how the composition of the tissue block, how the tissue was isolated, and the quality of fixation. On occasion when usable Gene Expression reads are lower with these samples, we were still able to get meaningful, biologically relevant results.
For best practices on maintaining RNA quality, see the resources listed below:
- Visium CytAssist Spatial Gene and Protein Expression Tissue Preparation Guide
- How should I fix and embed my tissue for the Visium CytAssist Spatial Gene and Protein assay?
- How should I store my FFPE tissue blocks?
- How do I transport my tissue slide to another facility when tissue has already been placed?
Products: Visium CytAssist Spatial Gene and Protein