Background: The dissemination of molecules through tissue establishes signaling gradients that drive complex developmental patterns, enables rapid responses to infection, and dictates the distribution of therapeutic drugs. However, our understanding of this molecular cartography would be vastly improved by charting molecular traffic throughout organ systems and cataloguing cell and tissue responses upon arrival and sensing. We recently developed “molecular tracking devices” — protein-DNA conjugates that are stable during tissue transit in vivo — to follow the distribution, acquisition, and retention of antigen in the immune system during an immune response. Molecular tracking devices enable previously inaccessible measurements of gene expression responses to signaling gradients and ligand-receptor interactions in living animals; here we expand this approach to study the dynamics and concentrations of tracking devices within tissue with the goal of identifying key gaps in the single-cell field and establish the commercial potential of molecular tracking devices.
Technical Innovation: The Dr. Tamburini and Dr. Hesselberth team have created DNA barcodes that can be linked to antigens or drugs of interest to track them within in-vivo systems. Using single-cell mRNA sequencing or spatial transcriptomic platforms, users can quantify both antigen/drug uptake and the length of time antigens/drugs are retained at the single cell level. To protect DNA barcode from exonuclease and endonuclease degradation, phosphorothioate linkages were incorporated into the oligonucleotide. Within murine models, the team was able to successfully track and quantify the uptake of a protein of interest (ovalbumin) into various cell types (Figure 1). The team found that the DNA tags successfully resisted degradation and did not impact immune response within their murine models. While ovalbumin was used in the team’s research, other antigens or drugs could be attached to the DNA tag to understand uptake and delivery within an in-vivo system. These quantitative analyses cannot be performed with currently available research tools.
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http://cuamc.technologypublisher.com/files/sites/cu5259h_ncs_update.pdf