BioMicroCenter:SpTx

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The BioMicro Center supports two complementary approaches to spatial transcriptomics: 10x Visium HD from 10x Genomics for slide-based spatial gene expression, and AVITI24 in situ sequencing from Element Biosciences. The choice of method depends on your tissue type, resolution requirements, and experimental goals. We strongly encourage consultation with BMC staff prior to beginning a spatial experiment.

10X VISIUM HD

10x Visium HD is a spatial transcriptomics platform that profiles whole-transcriptome gene expression across intact tissue sections at single-cell-scale resolution. Tissue sections are processed through the CytAssist instrument, which transfers RNA complexes onto a capture slide containing a continuous array of 3,000,000 spatially barcoded 2 µm squares covering a 6.5 × 6.5 mm capture area (~0.65 cm²). Two chemistries are available: a probe-based assay for human and mouse (FF, FxF, and FFPE), and a polyA-based assay (Visium HD 3') for fresh frozen tissue from any species. H&E or immunofluorescence (IF) staining can be performed prior to processing for protein co-detection.

ARRAY SPECIFICATIONS Capture area 6.5 × 6.5 mm (~0.65 cm²) per tissue Spots 3,000,000 barcoded squares Spot size 2 µm continuous (no gaps) Default analysis bins 8 µm and 16 µm (SpaceRanger); custom bins supported Tissues per slide 2 Multiomics H&E or IF staining prior to processing; antibody panels (FFPE, human only) CHEMISTRY OPTIONS Visium HD (probe-based) Visium HD 3' (polyA-based) Species Human, Mouse only Any species Tissue formats Fresh Frozen (FF), Fixed Frozen (FxF), FFPE Fresh Frozen (FF) only Notes ~3 probes per gene; exogenous sequences (GFP, reporters) require custom probes. A small fraction of genes are excluded by default due to predicted off-target activity: human excluded gene list (~2.1%), mouse excluded gene list (~1.8%). Species-agnostic; captures polyadenylated transcripts TISSUE PREPARATION REQUIREMENTS Fresh Frozen (FF) Fixed Frozen (FxF) FFPE Section thickness 10–20 µm 10 µm recommended 10–20 µm 5 µm Storage OCT, −80°C −80°C Room temp RNA quality DV200 ≥ 30% recommended; higher scores improve sensitivity Histology Sectioning & H&E performed by the Tang Histology Facility (KI). Coordinate with BMC staff to arrange. For current pricing, see BioMicroCenter:Pricing → Single Cell & Spatial Services → Visium. Sequencing is charged separately; 400–800M reads per tile recommended for Visium HD. SUBMISSION Submit via iLabs (assisted service). Coordinate with BMC staff at least one week in advance. Tissue processing begins at the Tang Histology Facility; please arrange sectioning separately.

10X Visium HD FAQs for Users 1) What tissues are compatible? Visium HD (probe-based) supports human and mouse tissues in FF, FxF, and FFPE formats. Visium HD 3' (polyA) supports fresh frozen tissue from any species. 10x Genomics maintains a list of tested tissues. Additional optimization may be required for tissues with high fat content. Consult BMC staff before starting. 2) How do I prepare and submit tissue? Tissue sectioning and H&E staining are performed by the Hope Babette Tang (1983) Histology Facility at the Koch Institute. Coordinate with both the Histology Facility and BMC staff before your first submission. FF tissue should be embedded in OCT and stored at −80°C; FFPE blocks at room temperature. 3) What is the resolution? Visium HD captures data at 2 µm bin resolution (single-cell scale). SpaceRanger aggregates into 8 µm and 16 µm bins for higher sensitivity by default; custom bin sizes are also supported. Cell segmentation based on H&E imaging is available in the Loupe Browser pipeline. 4) How much sequencing is needed? 400–800M reads per tile is recommended for Visium HD. Sequencing is submitted separately via iLabs. Compatible platforms include NovaSeq SP, NovaSeqX, and Element AVITI. Discuss read depth with BMC staff based on your tissue type and goals. 5) Does Visium HD require permeabilization optimization? No. Visium HD does not require permeabilization time optimization. For fresh frozen tissues new to the platform, a test array is available to verify tissue quality and placement before committing a full experiment. 6) Can I detect proteins alongside gene expression? Yes. H&E or IF staining can be performed prior to CytAssist processing. Antibody panels for protein co-detection (human tissues, FFPE only) are available from 10x Genomics. Discuss your experimental design with BMC staff in advance. 7) What analysis outputs are provided? The BMC delivers FASTQ files, aligned BAM files, the spatial feature-barcode matrix (raw and filtered at 2, 8, and 16 µm bins), and a Loupe Browser file. Downstream informatics support is available through BMC staff. 8) Can I combine Visium HD with single-cell data? Yes. Visium HD data integrates with single-cell or single-nucleus RNA-seq (e.g., 10x Chromium Flex) for cell-type deconvolution. Starting from the same tissue block, nuclei can be isolated for Flex single-cell gene expression. Coordinate with BMC staff if planning a multi-modal experiment. 9) My gene of interest is exogenous (GFP, reporter, viral) or missing from the probe set. What are my options? The probe-based Visium HD assay does not capture exogenous sequences by default. Custom probes can be designed for detection — see 10x Genomics technical note CG000621. Additionally, a small fraction of endogenous genes are excluded from the standard probe sets due to predicted off-target activity: see the human excluded gene list (~2.1% of genes) and mouse excluded gene list (~1.8% of genes). Alternatively, the polyA-based Visium HD 3' assay captures all polyadenylated transcripts and does not require custom probes.