The Visium Spatial Gene Expression Solution produces spatially barcoded, Illumina® sequencer-ready libraries.

Please note that Dual Index Kit TT, Set A (PN-1000215) is required for use with the Visium Spatial Gene Expression Solution.

Compatible Sequencers:

• Illumina® NovaSeq X Series
• Illumina® NovaSeq 6000
• Illumina® HiSeq 3000/4000
• Illumina® HiSeq 2500 (Rapid Run and High Output)
• Illumina® NextSeq 2000
• Illumina® NextSeq 500/550
• Illumina® MiSeq*
• Illumina® iSeq 100*

Dual-Indexed Sequencing Run: Visium Spatial Gene Expression libraries are dual-indexed. We do not recommend sequencing these libraries with a single-index configuration.

PhiX Spike-In Recommendations: 1%

* Recommended for library QC only

Visium Spatial Gene Expression Libraries

Minimum Sequencing Depth: We recommend a minimum of 50k read pairs per spot covered with tissue. To calculate this, first estimate the % of capture area covered by the tissue section based upon the H&E brightfield image. Then use the following formula to calculate the recommended sequencing depth:

• Total sequencing depth = (Coverage Area x total spots on the Capture Area) x 50,000 read pairs/spot

Example calculation for 60% tissue coverage: (0.60 x 5,000 total spots) x 50,000 read pairs/spot = 150 million total read pairs for that sample

Additional guidelines for estimating the number of spots covered: Determining number of spots covered with tissue may be performed visually or by using the Visium Manual Alignment Wizard in Loupe Browser for a more accurate measurement.

** Shorter reads than indicated above can lead to decreased application performance. Sample index reads must not be shorter than indicated. Any read can be longer than recommended.