Human Lung Cancer (FFPE)

10x Genomics obtained FFPE Human Lung Cancer tissue blocks from Avaden Biosciences. The tissue was sectioned as described in Visium CytAssist Spatial Gene Expression for FFPE – Tissue Preparation Guide Demonstrated Protocol (CG000518). Tissue sections of 5 µm was placed on a standard glass slide, then H&E-stained following deparaffinization. Sections were coverslipped with 85% glycerol, imaged, decoverslipped, followed by dehydration & decrosslinking Demonstrated Protocol (CG000520). The glass slide with tissue section was processed via Visium CytAssist instrument to transfer analytes to a Visium CytAssist Spatial Gene Expression slide. The probe extension and library construction steps follow the standard Visium for FFPE workflow outside of the instrument.

• Diagnosis:  Lung, Squamous Cell Carcinoma

The H&E image was acquired using Olympus VS200 Slide Scanning Microscope with these settings:

• Olympus Objective magnification: 20x UPLXAPO Objective
• Numerical Aperture: 0.8
• ScopeLED light source: Xcite Novum
• Camera: VS-264C
• Exposure: 500 µs

Libraries were prepared following the Visium CytAssist Spatial Gene Expression Reagent Kits for FFPE User Guide (CG000495).

• Sequencing instrument: Illumina NovaSeq, flow cell HNNNMDSX3 (lane 1-4)
• Sequencing depth: 35,785 reads per spot (sequencing saturation of 33%)
• Sequencing configuration: 28bp read 1 (16bp Visium spatial barcode, 12bp UMI), 90bp read 2 (transcript), 10bp i7 sample barcode and 10bp i5 sample barcode
• Dual-Index set: SI-TS-B1
• Slide: V42A20-354
• Area: D1

Key metrics were:

• Spots detected under tissue: 3,858
• Median genes per spot: 6,174
• Median UMI counts per spot: 13,991