Isothermally Amplified Proximity Ligation Assays for Biothreat Agents

Echo Technical and the Ellington Lab at the University of Texas at Austin have performed initial testing of a demonstration biosensor capable of using aptamer-based receptors (Biological Reconfigurable Interface Electronics For Classification and Analysis of Selected Elements, or BRIEFCASE). We propose to adapt a recently developed technology, proximity ligation assays, to a field-deployable device based on the BRIEFCASE to perform rapid, specific, hypersensitive detection and identification of the unique protein biosignatures of biothreat agents. To date, the proximity ligation assay has relied on polymerase chain reaction (PCR) for signal amplification. While PCR has previously been adapted to fielded devices, these devices have tended to be relatively bulky, in part because of the hardware required for thermal cycling. In order to simplify reagent addition and ultimately the device itself, we will therefore instead utilize isothermal amplification reactions that are as sensitive as PCR, but that require only a single addition of reagents in order to amplify nascent signals. This new technique, isothermally amplified proximity ligation assay (IAPLA), and an IAPLA-based biosensor prototype will be demonstrated to support goals identified for sensitivity, miniaturization, and field deployability, with feasibility demonstrated by the end of Phase I.