Biomaterials
A mouse lung (fresh frozen) was obtained from Charles River Laboratories.
- Strain: C57BL/6
- Sex: Male
- Age: 8 weeks
Sample preparation
A 10 µm section was taken with a cryostat (Epredia CryoStar NX70). Tissue block preparation, sectioning, H&E staining, and imaging followed the Visium HD Fresh Frozen Tissue Preparation Handbook (CG000763).
Imaging
- Image type: H&E
- Microscope: Olympus VS200 Slide Scanner
- Objective magnification: 20X
- Numerical Aperture: 0.8
- ScopeLED light source: VS200 LED, Olympus integrated bright field source
- Camera: iDS VS-264C, Olympus scanner integrated camera
- Exposure: 500 microseconds
Assay workflow
Probe hybridization, probe ligation, slide preparation, probe release, extension, and library construction followed the Visium HD Spatial Gene Expression Reagent Kits User Guide (CG000685).
- Slide serial number: H1-QD7CCD4
- Area: D-1
- Instrument: Visium CytAssist
- Probe set: Visium Mouse Transcriptome Probe Set v2.0
Sequencing
- Indexing: Dual index plate TS set A; sample index H5
- Sequencing instrument: Illumina NovaSeq 6000
- Sequencing configuration: 43 bp read 1, 50 bp read 2, 10 bp i7 sample index, and 10 bp i5 sample index
- Sequencing depth: 985.5 million read pairs (combined across two flow cells)
Analysis
Space Ranger v3.1.1 was used to map FASTQ files to the reference, detect tissue, align the data to the microscope and CytAssist images, and output feature-barcode matrices for further analysis.
How to view data
To get started, download Loupe Browser v8.0 or later to explore the Loupe file, or read more about the other Space Ranger outputs.