36C24224P1582_1.docx

Award Notice

GENERAL INFORMATION

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SET-ASIDE

Network Contracting Office 2 James J. Peters VA Medical Center 130 West Kingsbridge Road Bronx NY 10468-3904

POINT OF CONTACT*

Contract Specialist Paul Speedling Paul.Speedling@va.gov 914-737-4400 x202107

AWARD INFORMATION

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AWARDEE ADDRESS LINE 2*

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ADDITIONAL INFORMATION

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DESCRIPTION

STATEMENT OF WORK

10x Genomics Xenium Analyzer/10x Genomics Visium CytAssist

Background:

The goal of this project is to update the Single Cell Genomics Facility to assist Northport VA Medical Center and affiliated investigators in performing high throughput multiomic analysis at the single cell level. In particular, we will purchase and set up instrumentation that will allow us to evaluate the genomic, proteomic, and transcriptomic variations between individual cells maintaining the spatial information in the samples. This instrumentation will significantly enhance the ability of Northport VA scientists to investigate mechanisms of disease progression at the level of single cells in many tissues from clinical pathology slides as well as experimental animal models. The recent development of high throughput in situ based spatial -omics technologies allows the ability to identify the heterogeneity in gene and protein expression with subcellular resolution. Obtaining spatial information in addition to gene and protein expression in single cells is proving to be instrumental for unraveling the pathophysiological mechanisms underlying tumor progression and metastasis formation, immune responses to pathogens and virtually all the instances where crosstalk of different cell types is critical for the development and progression of the disease.

Objectives:

a) To establish a workflow that allows whole transcriptome spatial sequencing analysis of tissues processed with the Xenium Analyzer (10x Genomics). In particular, the Xenium (10x Genomics) will be used to process tissue slides obtained from the biological samples to tag molecules from individual cells and reveal expression and localization of transcripts and proteins in the samples at a subcellular resolution. Tissue sections processed using the Xenium In Situ workflow remain largely intact post-run and can be used for additional whole transcriptome spatial RNA sequencing. The latter application requires the Visium CytAssist instrument in order to transfer the tissue sample from the Xenium slide onto the specialized Visium slides containing the whole transcriptome probes for spatial gene expression analysis.

b) The data gathered with this workflow will allow the research projects of all the participating PIs to yield novel findings and to remain compelling and competitive for funding. These projects have direct relevance to Veterans’ health.

Performance Period:

30 days after award

Place of Performance:

Northport VAMC Research, Building 62, room 226 79 Middleville Rd Northport, NY, 11768-2200 Equipment will be tagged by Logistics and placed on Research EIL#408

Product Specifications: Research use only.

Features: The Visium CytAssist is a compact instrument designed to simplify spatial transcriptomics workflow by facilitating the transfer of transcriptomic probes from standard glass slides to 10x Genomics Xenium or Visium slides, enabling targeted or whole transcriptomic spatial profiling insights across entire tissue sections previously stored or freshly prepared, and expanded compatibility with FFPE and/or fresh frozen samples.

In the Visium CytAssist workflow, sectioning, tissue preparation, staining (H&E or IF), and imaging take place on a standard glass slide. Tissue samples previously placed on a Xenium slide can also be processed. After probe hybridization, two standard glass slides (or the Xenium slide) and a Visium slide with Capture Areas are placed in the CytAssist instrument so that the tissue sections on the standard slides can be aligned on top of the Capture Areas. Within the instrument, a brightfield image is captured to provide spatial orientation for data analysis, followed by hybridization of transcriptomic probes to the Visium slide. The remaining steps, starting with probe extension, follow the standard Visium spatial whole transcriptome workflow outside of the instrument.

Features: Xenium is an end-to-end platform from 10x Genomics that enables high-throughput subcellular mapping of 100s to 1,000s of RNA targets alongside multiplexed protein in the same tissue section. Compatible with fresh frozen (FF) and formalin-fixed, paraffin-embedded (FFPE) tissues, the platform includes the Xenium Analyzer, a versatile and easy-to-use instrument; sensitive and specific chemistry; a diverse menu of customizable panels; and software.

Data can be visualized with the built-in software suite Xenium Explorer or community-developed analysis tools. The Xenium platform has a robust roadmap to enhance the core platform with more capabilities and analytes.

The Xenium workflow starts with sectioning tissues onto a Xenium slide (Figure 1). The sections are then treated to preserve and access the RNA with circularizable DNA probes. The DNA probes are flanked by two regions that independently hybridize to the target RNA and also contain a gene-specific barcode sequence. Ligation of the probe ends to each other, then generates a circular DNA probe which is enzymatically amplified. In the event that one part of the probe experiences off-target binding, ligation will not occur, thus suppressing off-target signals and ensuring high specificity.

Multiplexed protein detection, a proven capability that will be enabled on the platform in the future, will leverage DNA-barcoded antibodies for labeling of protein targets in each tissue section. The Xenium workflow is also non-destructive. With the sample morphology intact at the end of the workflow, both hematoxylin & eosin (H&E) and immunofluorescence (IF) staining are possible on the same tissue section, enabling the direct comparison of Xenium transcriptional data to morphological data.

Security:

The C&A requirements do not apply, and a Security Accreditation Package is not required.

This contract does not involve access to VA sensitive information and does not involve access and/or connection to the VA information system.

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