Focus images on the MiniSeq and NextSeq 500/550

At the beginning of Read 1 on the MiniSeq and NextSeq 500/550, the instrument will determine the best Z position (distance between the objectives and flow cell) where the clusters are well-focused. During this process, the instrument will take and save a series of Focus Images. Many of the images will be completely back, though some will show the clusters (clusters are visible in the images when the Z position is correct, allowing for proper focus of the clusters on the flow cell).
Technical Support can evaluate Focus Images to diagnose clustering issues (failed to cluster/ underclustering and overclustering) or an unstable focus model.
  • Location of Focus Images:
    • MiniSeq: D:\Illumina\MiniSeq Sequencing Temp[Run Folder]\Images\Focus
    • NextSeq 500/550: D:\Illumina\NextSeq Control Software Temp[Run_Folder]\Images\Focus
Example Focus Images:
Example Image
Normal cluster density ~220 k/mm2
Standard clustering
rtaImage (1).jfif
Ultra low cluster density
All cameras disabled at cycle 1 Visible background between clusters White edges due to increased intensity
Ultra Low Density.jfif
Very low cluster density
All cameras disabled at cycle 1 Visible background between clusters
Very low cluster density.jfif
Low cluster density
Low intensity or high G base calling Visible background between clusters No white edges
Low cluster density.jfif
High cluster density
No white edges Visible fold-like artifacts
High cluster density 1.jfif
High cluster density 2.jfif
No clusters
White edges indicate increased intensity to compensate for initial lack of clusters
No clusters.jfif
For any feedback or questions regarding this article (Illumina Knowledge Article #6325), contact Illumina Technical Support [email protected].
Last modified 23d ago
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