Question: Why did my Nuclei Isolation Column clog during centrifugation?
Answer: When isolating nuclei using the Chromium Nuclei Isolation Kit, there are several potential causes that the Nuclei Isolation Column will clog.
(A) Column with successful flowthrough and (B) Clogged column
The most likely potential causes for the column to clog are:
- The mass of tissue used during isolation was greater than 50mg
- The tissue was not fully dissociated
- The lysis time was not sufficient for full dissociation
- The tissue type is not compatible with the Chromium Nuclei Isolation Kit
The following are solutions to mitigate the column from clogging:
- Ensure the tissue is not on the list of incompatible tissue types
- Ensure the tissue mass is within the recommended tissue range of 3-50mg
- Increase the number of pestle strokes for complete tissue dissociation
- Lyse tissue for a longer period of time
To recover a sample that clogs a Nuclei Isolation Column, transfer liquid remaining on top of the spin column to a new Nuclei Isolation Column leaving behind any debris stuck to the column filter. Place the column into the same Collection Tube and repeat the centrifugation step. Contact 10x Genomics Technical Support at 10xgenomics.com for further assistance.
Products: Single Cell Gene Expression, Single Cell Immune Profiling, Single Cell ATAC, Single Cell Multiome ATAC + GEX, Fixed RNA Profiling