This presentation by Dr. Angela Dorigatti of Takara / Curio Biosciences introduces Trekker, a novel spatial transcriptomics technology designed to achieve true single-cell resolution. Dr. Dorigatti begins by outlining the limitations of existing methods: sequencing-based assays like Visium suffer from low gene sensitivity and mixed-cell data spots that require deconvolution, while microscopy-based methods are limited in the number of targets.
The Trekker technology addresses these issues by reversing the capture process. Instead of releasing mRNA onto a slide, Trekker uses a UV-activated slide to release spatially barcoded oligonucleotides up into a fresh-frozen tissue section, directly tagging individual nuclei. These barcoded nuclei are then isolated and processed through standard single-cell workflows (e.g., 10x Genomics, BD Rhapsody), generating both a gene expression library and a spatial location library. This approach maintains the high sensitivity of single-cell sequencing (hundreds to thousands of genes per cell) and determines cell location with two-micron accuracy through a triangulation of barcode ratios. Crucially, this workflow re-enables multi-omic applications like ATAC-seq and TCR/BCR analysis in a spatial context. The presentation also covers practical aspects, bioinformatics solutions via partnerships with Latch Bio and Bio-Turing, and confirms an FFPE-compatible version (Trekker FX) is in development.
0:35 Introduction to Technology: Dr. Angela Dorigatti introduces Takara Bio's Seeker (spatial RNA-seq mapping) and Trekker (single-cell spatial mapping) technologies, born from the pioneers behind Drop-seq and CITE-seq.
3:09 The Problem with Current Methods: Existing spatial technologies face a trade-off. Sequencing-based methods like Visium capture only 10-15% of mRNA and mix signals from multiple cells into one spot, requiring complex deconvolution. Microscopy-based methods are limited by the number of probes.
5:54 Trekker's Single-Cell Solution: Based on the "slide-tags" paper, Trekker adds a spatial barcode to each nucleus before it enters a standard single-cell sequencing workflow, thereby preserving true single-cell resolution and sensitivity.
6:34 How Trekker Works: A fresh-frozen tissue section is placed on a tile coated with beads, each carrying a unique spatial barcode. A UV light releases these barcoded oligos, which diffuse up into the tissue and tag the individual nuclei within.
7:42 Integration with Standard Workflows: After tagging, the nuclei are isolated into a suspension and can be used as direct input for common platforms like 10x Genomics or BD Rhapsody. The process generates two distinct libraries: one for gene expression and one for spatial location.
9:53 High-Resolution Triangulation: A cell's precise location (within 2 microns) is calculated based on the ratio of barcodes it receives from surrounding beads, similar to GPS triangulation. This process also allows for confident identification and removal of doublet artifacts.
14:33 Re-enabling Multi-Omics: A key advantage of the Trekker workflow is its ability to integrate multi-omic assays that were lost in traditional slide-based spatial methods, including ATAC-seq (chromatin accessibility) and TCR/BCR sequencing.
17:07 Superior Data Quality: Unlike slide-capture methods that produce noisy data and blended cell clusters, Trekker provides high-sensitivity data (hundreds to thousands of genes per cell), resulting in distinct, easy-to-interpret cell clusters.
18:43 Efficient Cell Sampling: Because Trekker analyzes a thin tissue slice, researchers can achieve high-density spatial maps by sequencing a fraction (e.g., 20-30%) of the total nuclei isolated, making experiments more cost-effective.
20:53 Multi-Modal Case Study: A metastatic melanoma sample was successfully analyzed using Trekker to generate three datasets from the same tissue: single-nuclei RNA-seq, ATAC-seq, and TCR enrichment data.
22:05 User-Friendly Bioinformatics: Data analysis is supported through a local pipeline or cloud-based platforms like Latch Bio and Bio-Turing, which offer user-friendly, point-and-click interfaces for complex spatial analysis.
27:37 Low Sequencing Requirement: The spatial barcode library requires very little sequencing depth compared to the gene expression library—only 5,000 reads per nucleus for 10x Genomics and 1,000 for BD Rhapsody.
30:27 FFPE Version Coming Soon: An FFPE-compatible version, named Trekker FX, is planned for release by the end of the year. It will integrate with the 10x Genomics Flex kit and allow for H&E imaging on the same tissue section.