Probes and cell arrays for detection of bacterial toxins

DESCRIPTION (provided by applicant): Microarray technology offers an opportunity for simultaneous monitoring of the behavior of multiple markers within a mammalian cell and ultimately could be used for detection and elucidation of mechanisms of action of different biologically active agents, including those that are considered a threat in the biodefense area. The "predictive power" of probes used in diagnostic arrays and the cost of their application will be crucial parameters for introduction of array technology into routine diagnostic practices. We propose to develop an innovative robust system that will allow for the identification and isolation of the specific human DNA sequences that are up or down regulated in response to interaction between a eukaryotic cell with specific biologically active agents or groups of biologically active agents. Such sequences could be used as the basis for the construction of a cell array based detection system. In Phase I, we will demonstrate the technical feasibility of all fundamental aspects of the application for elaboration during subsequent phases. Our approach will be to use a flow cytometer cell sorting process to identify and isolate genetic elements, previously fused with the reporter sequences, which respond to the specific toxic agent challenge. In Phase II, our principal effort will be to extend our library of responsive elements that will be used for detection of major toxic agents and construct a prototype of the diagnostic cell array. The ultimate goal of this research will be to provide a general and robust approach for the detection of biologically active agents, especially when these agents have been engineered to elude immunoassays which are currently available. Cell arrays offer a new opportunity for sensitive and precise monitoring of biologically active substances. The goal of this application is to develop a system for identification of regulatory elements that will allow a substantial extension of discriminative abilities of cell arrays and creation of cell arrays capable of detection and identification of potential biowarfare agents.