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Visium CytAssist Gene and Protein Expression Library of Human Breast Cancer, IF, 6.5mm (FFPE)

CytAssist Spatial Gene and Protein Expression dataset analyzed using Space Ranger 2.1.0

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10x Genomics obtained FFPE Human Breast Cancer tissue block from Avaden Biosciences. The tissue was sectioned as described in Visium CytAssist Spatial Gene and Protein Expression for FFPE – Tissue Preparation Guide (CG000660). 5 µm tissue sections were placed on Superfrost glass slides, deparaffinized, decrosslinked, IF stained, and coverslipped. Sections were imaged and decoverslipped, as per the Demonstrated Protocol (CG000659).

Sample Description:

  • Sex: Female
  • Age: 50s
  • Date Excised: 2007
  • DV200: 56.0

Antibodies used for immunofluorescence (IF) staining:

  • Channel 1 = 1:200 DAPI (Thermo Fisher, Cat. No. 62248)
  • Channel 2 = 1:100 Vimentin, 594 nm (Biolegend Cat. No. 677804)
  • Channel 3 = 1:100 PCNA, 647 nm (Biolegend, Cat. No.307912)
  • Blocking buffer: 1X Blocking Buffer (10x Genomics Cat. No. 2000554); 2 U/uL RNase Inhibitor (10x Genomics Cat. No. 2000556)

The IF image was acquired using Olympus Slide Scanning Microscope with these settings:

  • Olympus Objective Magnification: 20X UPLXAPO Objective
  • Numerical Aperture: 0.8
  • ScopeLED light source: Xcite Novum
  • Camera: Hamamatsu Orca FusionBT sCMOS
  • The IF image was upscaled for aesthetic purposes

Libraries were prepared following the Visium CytAssist Spatial Gene and Protein Expression Reagent Kits User Guide (CG000494):

  • Sequencing instrument: Illumina NovaSeq, flow cell H3TVGDSX7 (lane 4)
  • Sequencing depth (Gene): 35,000 reads per spot
  • Sequencing depth (Protein): 20,000 reads per spot
  • Sequencing configuration: 28bp read 1 (16bp Visium spatial barcode, 12bp UMI), 50bp read2 (probe insert/antibody barcode), 10bp i7 sample barcode and 10bp i5 sample barcode
  • Dual-Index set: SI-TS-D11, SI-NT-D11
  • Slide: V43J24-072
  • Area: A1

Key metrics were:

  • Spots detected under tissue: 4,169
  • Median genes per spot: 5,473
  • Mean reads per spot: 44,009
  • Mean Antibody reads per spot: 43,038

This dataset is licensed under the Creative Commons Attribution license.