Adult Mouse Brain, IF Stained (FFPE)

10x Genomics obtained FFPE mouse brain tissue from Adlego and embedded by BioIVT. The tissue was sectioned as described in Visium Spatial Gene Expression for FFPE – Tissue Preparation Guide Demonstrated Protocol (CG000408). Tissue sections of 5 µm were placed on Visium Gene Expression slides, then stained following Deparaffinization, Decrosslinking, Immunofluorescence Staining & Imaging Demonstrated Protocol (CG000410).

• Sex: Male
• Age: >8 weeks
• Strain: C57BL/6
• Section Orientation: Coronal

Antibodies used for Immunofluorescence staining:

• Channel 1 = 1:100 NeuN (Abcam, Cat. No. ab190565)
• Channel 2 = 1:200 GFAP (Sigma-Aldrich, Cat. No. C9205-.2ML)
• Channel 3 = 1:5000 DAPI (Thermo Scientific, Cat. No. 62248)
• Blocking buffer: 1X PBS; 2 % BSA; 0.1 % Tween 20; 1 U/uL Roche Protector RNase Inhibitor (based on CoA)

The IF image was acquired using Metafer Slide Scanning Microscope from MetaSystems with these settings:

• Zeiss Plan-Apochromat 20x objective
• Numerical Aperture: 0.8
• Zeiss Colibri 7 LED light source
• MetaSystems CoolCube 4c camera

Libraries were prepared following the Visium Spatial Gene Expression Reagent Kits for FFPE User Guide (CG000407 Rev A).

• Sequencing instrument: Illumina NovaSeq, flow cell HCYTVDSX2 (lane 1-2)
• Sequencing depth: 30,337 reads per spot
• Sequencing configuration: 28bp read 1 (16bp Visium spatial barcode, 12bp UMI), 120bp read2 (transcript), 10bp i7 sample barcode and 10bp i5 sample barcode
• Dual-Index set: SI-TS-A9
• Slide: V11J26-128
• Area: A1

Key metrics were:

• Spots detected under tissue: 2,438
• Median genes per spot: 6,056
• Median UMI Counts per spot: 17,096

This dataset is licensed under the Creative Commons Attribution 4.0 International (CC BY 4.0) license. 10x citation guidelines available here.