Use of nanotechnology to rapidly detect human pathogens

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DESCRIPTION (provided by applicant): In order to reduce escalating health care costs, rapid "point-of-care" (POC) testing has dramatically grown over the last decade. Due to their relative ease, immunologic approaches are frequently used for such tests. Molecular analysis (i.e., based upon DNA and/or RNA), however, has largely been limited to the clinical laboratory setting, where the inherently more complicated sample preparation and analysis procedures can be performed. Nanotechnology provides a unique opportunity to bring molecular technology to the patient, thus improving diagnostic data and further reducing healthcare costs. We propose to develop a nanomedical device for the direct detection of specific nucleic acid sequences in human pathogens. By eliminating enzymatic gene amplification steps, POC testing can be more easily accomplished within the rapid time frame expected for such tests. The detection of pathogenic organisms will be accomplished by detecting abundant rRNA sequences. Two types of oligonucleotide probes will be designed and utilized. One ("capture" probe) will be bound to a magnetic bead that will serve to bind to and immobilize the target sequences within the proposed device. The second oligonucleotide probe ("reporter" probe) will be bound to liposome nanovesicles, which in turn, will encapsulate quantum dots (QDots). Because QDots can be "tuned" to fluoresce at specific wavelengths depending on their size, this feature will permit us to ultimately design a simple multianalyte test during Phase II. The target rRNA sequences of the pathogenic organism will be selected by first predicting their secondary structure (using free energy calculations) and then evaluating those sequences for specificity and availability for hybridization. The "capture" and "reporter" probes will be designed to hybridize to these available sequences. Integration of these technologies into a nanomedical diagnostic device will permit the rapid detection of specific nucleic acid sequences without the need for prior gene amplification, thus improving the speed and ease-of-use of molecular diagnostic assays in the POC and other human in vitro diagnostic marketplaces. Finally, the successful development of a "sample in - results out" device whereby no sample manipulation is required, will dramatically simplify the performance of such tests.