Question: Can RNase inhibitors be used in the preparation of my sample?
Answer: For preparing single cell suspensions, we have not tested using RNase inhibitors in the cell washing or resuspension buffers. However, we do recommend using RNase inhibitors when preparing nuclei suspensions and methanol-fixed cells, as detailed in our Demonstrated Protocols for Isolation of Nuclei for Single Cell RNA Sequencing and Methanol Fixation of Cells for Single Cell RNA Sequencing:
- Isolation of Nuclei for Single Cell RNA Sequencing
In this Demonstrated Protocol, the Nuclei Washing and Resuspension Buffer contains 0.2U/µl RNase Inhibitor (Sigma Aldrich PN-3335399001). We have tested RNase Inhibitor concentrations up to 1.0 U/µl and did not see degradation in 10x assay performance.
- Methanol Fixation of Cells for Single Cell RNA Sequencing
In this Demonstrated Protocol, the Rehydration Buffer contains 0.5U/µl RNase Inhibitor (Sigma Aldrich PN-3335399001).
Note: RNase Inhibitor in the Nuclei Washing and Resuspension Buffer or Rehydration Buffer may result in a translucent/opaque rather than clear aqueous phase after breaking the GEMs with Recovery Agent. This is normal and will not impact downstream operations.
Products: Single Cell 3', VDJ