A MICROFLUORIMETRIC DNA HYBRIDIZATION ASSAY

THE GOAL OF THIS RESEARCH IS TO DEVELOP A RAPID AND HIGHLY SENSITIVE DIRECT FLUORESCENT DETECTION SYSTEM FOR DNA HYBRIDS. DNA HYBRIDIZATION PROBES ARE USEFUL FOR DETECTING SPECIFIC PATHOGENS AND GENETIC ALTERATIONS. RADIOAUTOGRAPHY, CURRENTLY THE MOST SENSITIVE DETECTION TECHNIQUE, CAN VISUALIZE 15 PICOGRAMS OF TARGET DNA IN AN OVERNIGHT EXPOSURE. IN ORDER TO FULLY UTILIZE THE POTENTIAL OF DNA PROBES IN A CLINICAL SETTING, A SIMPLE, RAPID, SENSITIVE, NONRADIOACTIVE DETECTION METHOD IS NEEDED. FLUORESCENCE DETECTION, WITH ITS HIGH INHERENT SENSITIVITY, SHOULD PROVIDE SUCH A SYSTEM. IN PHASE I WE PROPOSE TO INVESTIGATE A FLUORESCENT HYBRID DETECTION SYSTEM UTILIZING FLUORESCENT OLIGONUCLEOTIDE HYBRIDIZATION PROBES IN A FOCUSED HYBRIDIZATION FORMAT, IN COMBINATION WITH A HIGHLY SENSITIVE MICROFLUORIMETER. PRELIMINARY RESULTS SUGGEST THAT INSTANTANEOUS DETECTION LIMITS FOR SUCH A SYSTEM ARE AT A LEVEL PROMISING SUPERIOR PERFORMANCE AND UTILITY. IN PHASE II WE PROPOSE TO CONDUCT TRIALS OF THE SYSTEM FOR DETECTING VIRAL AND BACTERIAL DNA TARGETS IN CLINICAL SAMPLES, WITH THE GOAL OF PRODUCING A RAPID, SIMPLE, AND SPECIFIC DETECTION SYSTEM.