Troubleshooting Regions of Reduced %Passing Filter on NovaSeq X Series

Background

The NovaSeq X Series uses a highly sensitive optics system to enable rapid flow cell scanning. Small debris under the flow cell surface can result in deflections of the flow cell surface and cause localized regions with reduced clusters passing filter or quality scores, similar to the following:

%Passing Filter (%PF) and Quality Scores (% >= Q30)

Resolution Actions

To mitigate this issue, Illumina recommends using using Polynit Heatseal Wipe ( VWR, catalog #68310-176, or equivalent general lab supplier) and 70% isopropyl alcohol to clean the underside of the flow cell and the top of the flow cell holder for every sequencing run. Previous guidance recommended cleaning when debris was identified, and this is now updated to recommend cleaning for each sequencing run setup.

Do NOT use other lint-free wipes (ex. KIM wipes or lens paper) as this can result in debris remaining on the flow cell and increase the risk of this phenotype.

When cleaning the flow cell and flow cell holder, wipe lengthwise up and down the length of the flow cell and flow cell holder. Do NOT wipe side-to-side as this can result in debris from the wipe depositing on the flow cell or flow cell holder.

For the flow cell, do not touch the gaskets at the top and bottom of the flow cell. For the flow cell holder, do not wipe debris into the line of vacuum ports at the middle of the flow cell holder.

For any feedback or questions regarding this article (Illumina Knowledge Article #9570), contact Illumina Technical Support [e-mail].