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

Please note that Dual Index Kit TS, Set A (PN-1000251) is required for use with the Visium HD Spatial Gene Expression Solution.

Compatible Sequencers:

• Illumina® NovaSeq 6000
• Illumina® NextSeq 2000
• Illumina® NovaSeq X Series

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

PhiX Spike-In Recommendations: 1%

Visium HD Spatial Gene Expression Libraries

Minimum Sequencing Depth: The minimum sequencing depth for Visium HD is 275 million read pairs per fully-covered Capture Area. The recommended minimum sequencing depth was chosen because it achieved >50% sequencing saturation for >50% formalin fixed, paraffin embedded (FFPE) tissues tested at 10x Genomics. To achieve >50% saturation for >50% of fresh frozen (FF) tissues tested by 10x Genomics, 700 million read pairs per fully-covered Capture Area were required. To achieve >50% saturation for >50% of the Fixed Frozen (FxF) tissues tested by 10x Genomics, 500 million read pairs per fully-covered Capture Area were required. Please refer to the Visium HD Spatial Gene Expression Reagent Kits User Guide (CG000685) “Sequencing Depth” section for additional information.

To calculate this, first estimate the % of capture area covered by the tissue section based upon the CytAssist image. Then use the following formula to calculate the recommended sequencing depth:

• Minimum sequencing depth = (275 million read pairs * fraction of capture area covered with tissue)

Example calculation for 60% tissue coverage: (275,000,000 * 0.60) = 165 million total read pairs for that sample

** Shorter reads than indicated above can lead to decreased application performance.