Amplified Biothreat Signature Detection Microarray

The proposed work will exploit the first-stage enzymatic plus second-stage electrochemical signal amplification method developed in Phase I for highly sensitive detection of biothreat agents. An amplification of 1,200,000 per reporter enzyme molecule was achieved experimentally in Phase I. Because several molecules of the reporter enzyme will link to most biothreat agents, the overall amplification will be even larger per captured agent. Assay specificity will be achieved via synthetic globular DNA affinity recognition elements selected for binding pathogens, toxins, or other protein-based biothreats to enable their detection in the field. In Phase II, a complete biothreat microchip system will be developed and tested. Spatial resolution of the microchip array elements will be provided by using MEMS (microelectro-mechanical systems) technology to isolate each element in its own microcuvette, each containing microelectrodes. The chip will contain microfluidics capable of flowing the sample, wash, and reagent liquids through all microcuvettes, and the electrical outputs will provide unique biosignatures characteristic of specific threat agents. A small field-portable instrument to control microchip sample acquisition and provide signal handling and readout will be designed and built. An appropriate chip "package" will be designed to protect the microchip, enable sampling, and provide an interface with the instrument. The complete system will be evaluated under conditions suitable for field deployment using a biothreat surrogate.