Visium HD Spatial Gene Expression Library, Human Tonsil, IF (Fresh Frozen)

Biomaterials

A human tonsil (fresh frozen) with Reactive Follicular Hyperplasia was obtained from BioIVT.

• Sex: Male
• Age: 21 years

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: IF
• Microscope: Olympus VS200 Slide Scanner
• Objective magnification: 20X
• Numerical Aperture: 0.8
• ScopeLED light source: Excelitas X-Cite NOVEM-L
• Camera: Hamamatsu Orca FusionBT sCMOS (2304 x 2304 pixels (16bits), 6.5um pixel size, 95% QE, 89.1 fr/sec, ~1Gb/sec)
• Antibodies: CD20_488, aSMA_594, CD45_647
• Exposure: DAPI (ex 385 nm, em 447 nm): 500 microseconds; 488 (ex.475 nm, em 519nm): 150 milliseconds; 590 (ex 586 nm, em 628 nm): 20 milliseconds; 647 (ex 635nm, em 668nm): 125 milliseconds

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-MFH8V29
• Area: A1
• Instrument: Visium CytAssist
• Probe set: Visium Human Transcriptome Probe Set v2.0

Sequencing

• Indexing: Dual index plate TS set A; sample index G11
• 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: 1271.6 million read pairs

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.

This dataset is licensed under the Creative Commons Attribution license.