Questions & Answers
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SupportVisium
General
- Are the Visium slides charged?
- Are the Visium Spatial Tissue Optimization or Gene Expression slides re-usable if not all sections are used?
- Can I modify the Visium for fresh-frozen Spatial Gene Expression solution to be targeted?
- Can I run different tissue types on a single Visium Spatial Gene Expression Slide for the Visium for fresh-frozen assay?
- Can I use the Visium Spatial Slide Reset Demonstrated Protocol to practice section placement for Visium?
- Do the spatial barcodes differ across individual capture areas on the Visium Spatial Gene Expression Slide?
Visium for FFPE - General
- My thermal cycler lid temperature doesn't go lower than 50°C, what should I do?
- Can I perform Spatial Gene Expression analysis using older FFPE blocks?
- What is the purpose of the Tissue Adhesion Test and how should it be performed?
- What is the difference between formaldehyde, formalin, and paraformaldehyde?
- What fixation and embedding methods are compatible with Visium for FFPE?
- What sample handling methods can be implemented to minimize the likelihood of FFPE tissue detachment on Visium Slides?
Sample Preparation
- Can OCT affect bulk RNA extraction and RIN assessment?
- How can I more easily visualize the fiducial frame during tissue section placement?
- Can I section from the same tissue block across multiple days?
- Can OCT cover the Fiducial Frame or extend outside the capture area?
- Do I need to decontaminate the cryostat between different tissues types?
- How can I avoid overdrying of my tissue sections?
Visium for FFPE - Sample Preparation
- Why are there so many drying steps in the Visium for FFPE Demonstrated Protocols?
- Should I punch and re-embed my FFPE block so my tissue can fit within the Capture Area?
- Can I place FFPE sections onto a Visium Spatial Gene Expression Slide during multiple sectioning sessions?
- What should I do if my FFPE tissue has detached from the Visium Gene Expression Slide?
- How well does tissue adhere to Visium Slides with Visium for FFPE compared to other tissue-based assays?
- I don't have a desiccator, what can I use instead?
Tissue Fixation and Staining
- I have already optimized my antibodies for immunofluorescence on its own. Do I need to re-optimize my antibodies for the Visium for fresh-frozen Immunofluorescence assay?
- How much time does the Immunofluorescence Staining Demonstrated Protocol for Visium for fresh-frozen require?
- Does the Visium for fresh-frozen Immunofluorescence Demonstrated Protocol differ from standard immunofluorescence protocols?
- Is my antibody compatible with the Visium for fresh-frozen Immunofluorescence (IF) Staining Demonstrated Protocol?
- What is the maximum number of antibodies that can be used when performing Visium with Immunofluorescence?
- What tissue types have been validated with the Visium for fresh-frozen Immunofluorescence Demonstrated Protocol?
Visium for FFPE - Staining
- Is Visium for FFPE compatible with immunohistochemistry (IHC)?
- Can I use xylene substitutes during deparaffinization?
- How long is my FFPE tissue stable after coverslip removal?
- Can immunofluorescence staining and imaging have a negative impact on Visium for FFPE assay sensitivity?
- Does the Visium for FFPE Immunofluorescence Demonstrated Protocol differ from standard immunofluorescence protocols?
- What reagents are needed to perform the Visium for FFPE IF Staining Demonstrated Protocol?
Imaging
- Does the etched frame on the Tissue Optimization slide need to be in-focus to interpret the experimental results?
- Can Visium Gene Expression Slides be stored post H&E staining?
- Does using a coverslip during imaging affect Visium for fresh-frozen data quality?
- What are the imaging system requirements for running Visium for fresh-frozen?
- Is a color camera required for Hematoxylin & Eosin (H&E) imaging, or can a monochrome camera be used?
- What is the purpose of the Visium Spatial Imaging Test Slide for the Visium for fresh-frozen assay?
Visium for FFPE - Imaging
cDNA Quantification & Amplification
- Can I use an alternative to the KAPA SYBR Fast qPCR Master Mix (KK4600) for Cycle Number Determination during the Visium Gene Expression workflows?
- Can I use alternative methods to qPCR such as Nanodrop or Qubit for cDNA quantification during the Visium Gene Expression workflow?
- Which qPCR systems can be used for the Visium assay?
Visium for FFPE - Protocol Steps
- Can I use an alternative to the KAPA SYBR Fast qPCR Master Mix (KK4600) for Cycle Number Determination during the Visium and Visium for FFPE Gene Expression workflows?
- During qPCR for Cycle Number Determination of the Visium and Visium for FFPE Gene Expression workflows, what should the real time controller lid temperature be set to?
- Can I use the Dual Index Kit Plate TT Set A, 96 rxns (PN-1000215) for Sample Index PCR with Visium for FFPE?
- What does a good Visium Spatial Gene Expression for FFPE library look like?
- What is the structure of the final Visium for FFPE library?
- What Dual Index Plate is compatible with Visium for FFPE?
Library Construction
- Why were the Sample Index PCR conditions updated for Visium Spatial Gene Expression library preparation?
- What is the structure of the final Visium for fresh-frozen library?
- What does a good Visium Spatial Gene Expression for fresh-frozen library look like?
- Can I use the Chromium i7 Multiplex Plate (PN-2201013) for Sample Index PCR with Visium?
- What sample index plate is required for performing Sample Index PCR with Visium for fresh-frozen?
Visium for FFPE - Transcriptome Probe Panels
- Can I use PDX/xenograft tissue with Visium for FFPE?
- Can library molecules be generated from Probe Hybridization events with genomic DNA?
- If there is a mutation within the binding regions of two probes for our Visium for FFPE assay, will the probes still bind to the mRNA?
- What genes will not be detected by the Transcriptome Probe Panels?
- How were the Transcriptome Probe panels designed?
- How were the genes included in the Visium for FFPE human probe set chosen?
Sequencing
- Why do a fraction of my Visium reads contain the Template Switch Oligo (TSO) at the beginning of Read 2?
- Where can I find the Dual Index Kit TT Set A (PN-1000215) sample index sequences?
- Can Visium for fresh-frozen libraries be pooled with other 10x libraries?
- What sequencing parameters should be used for Visium for fresh-frozen libraries?
- What are the minimum recommended sequencing specifications for Visium for fresh-frozen libraries?
- Can I change the sequencing read lengths of the i5 and i7 indices?
Visium for FFPE - Sequencing
- Can Visium for FFPE libraries be pooled with other dual indexed 10x libraries?
- Where can I find the Dual Index Kit TS Set A sample index sequences?
- What is the suggested minimum sequencing depth for Visium for FFPE libraries?
- What sequencers are Visium for FFPE libraries compatible with?
- What are the recommended sequencing specifications for Visium for FFPE libraries?
Software
- How do I know if I used the correct image?
- How can I change spaceranger filtering for FFPE probes?
- Is my gene in the Visium for FFPE probe set?
- Do misprinted fiducial frame spots affect the performance of automated image alignment?
- The fiducials are misaligned in my web summary — how can I fix this?
- How can I transfer pathologist annotations to Loupe Browser?