TempO-LINC high throughput high sensitivity single cell gene expression profiling assay Ph II - Summary/Abstract:
Single cell gene expression assays have become critical tools for identifying functional subtypes of cells,
as well as changes within specific subtypes resulting from development, disease or therapy treatment. However,
most current methods are expensive, low in sample throughput and rely on oligo-dT primed reverse transcription.
Furthermore, there is no method to-date that enables investigators to consistently measure low expressed genes
from single cells, which prevents measurements of many key biomarkers and important genes and molecular
pathways. Another problem with existing single-cell sequencing methods is that many of the cell-associated
reads either fail to map to transcripts or map to uninformative ribosomal and mitochondrial genes. The
aforementioned limitations have all contributed to most single-cell sequencing studies being restricted to
relatively small numbers of samples and have prevented their wider scale adoption in biopharma, clinical,
translational and applied markets.
In Phase 1 SBIR development, we implemented a novel combinatorial split-pool-based workflow,
TempO-LINC, that adds cell-identifying molecular barcodes onto high-sensitivity gene expression probes. All
probes within the same cell receive an identical barcode, enabling the reconstruction of single-cell gene
expression (and protein) profiles across tens of thousands of cells. TempO-LINC is an instrument free approach
that multiplexes 96 1,000 cell samples per run, has a simple workflow that is amenable to automation and has
the highest gene detection rate of any commercial single-cell platform. Critically, TempO-LINC reduces both the
per sample assay cost as well as associated sequencing costs; furthermore, although we show the assay
simultaneously profiles the whole transcriptome (23,000 transcripts), it can also be targeted to measure only
genes/pathways of interest – dramatically reducing sequencing costs yet further. All of these features position
TempO-LINC as a potential disruptive technology that researchers, clinicians and drug developers can use for
new large-scale applications/studies using single-cell sequencing across thousands of samples while obtaining
improved data quality.
Our objective in this Phase 2 proposal is to take the basic TempO-LINC platform that we are currently
running and ready it for commercialization by optimizing the sensitivity, workflow, multi-omic capability and
software. To demonstrate the unique capabilities of TempO-LINC, we will benchmark it against 10X Chromium
data using PBMCs and demonstrate a high-impact application of our technology with our collaborators/partners.
Our approach relies on existing commercial TempO-Seq assays/technology, Phase 1 data that far exceeded all
milestones, and a solid foundation of intellectual property. BioSpyder’s TempO-LINC constitutes a powerful
single-cell platform with high potential for technical success, which will fill a critical need in the marketplace.
