Enzymatically amplified surface plasmon resonance imaging method using RNase H and RNA microarrays for the ultrasensitive detection of nucleic acids

A novel surface enzymatic amplification method that utilizes RNA microarrays in conjunction with the enzyme RNase H is developed for the ultrasensitve detection and analysis of target DNA molecules. The enzyme RNase H is shown to selectively and repeatedly destroy RNA from RNA-DNA heteroduplexes on gold surfaces; when used in conjunction with the label-free technique of surface plasmon resonance imaging, multiple DNA targets can be detected at a concentration of 10 fM on a single chip. In addition, this method is utilized for the sequence-specific detection of the TSPY gene in both purified and unpurified PCR products. Finally, in a series of kinetics measurements, the initial rate of hydrolysis is shown to depend directly on the surface concentration of DNA-RNA heteroduplexes.