Single-Molecule Spectroscopy for Nucleic Acid Analysis: A New Approach for Disease Detection and Genomic Analysis

Abstract

Recently developed single-molecule spectroscopy (SMS) permits the analysis of fluorescent mixtures one molecule at a time. SMS methods provide the means to make rapid measurements on small, complex samples without the need for separations and target amplification enabling a new class of ultrasensitive nucleic acid assays. Here we give a brief overview of the current state of the art of SMS nucleic acid analysis and discuss ongoing work in our laboratory on two-color single-molecule fluorescence detection of specific nucleic acid sequences. In the future, two-color SMS nucleic acid assays will be used for a variety of applications including: gene expression analysis, disease detection and genomics.

Single-Molecule Detection and Probe Strategies for Rapid and Ultrasensitive Genomic Detection

“…Over the past two decades a variety of optical SMD techniques (in solutions or on a solid surface) have been developed [1][2][3][4][5][6][7][8][9], greatly advancing fundamental bio-chemical and bio-physical studies. In-solution SMD is mainly based on the use of confocal fluorescence spectroscopy, which detects the emission from a single fluorescent molecule when it traverses through a femtoliter-sized detection volume.…”

“…The small detection volume of SMD, however, demands an efficient scheme for transporting molecules through the detection volume to improve the mass detection efficiency. 6 6 0 6 0 r R R E + = Recent advances in micro-or nanofluidics technologies have facilitated the controlled transport of minute amounts of fluids as well as the precise manipulation of biomolecules using electrokinetic [14,[21][22][23][24] or hydrodynamic forces [25,26]. With the high detection sensitivity of SMD and the high mass detection efficiency enabled by the use of microfluidic devices, the integrative microfluidic-SMD approach makes possible the accurate and quantitative analysis of nucleic acids at low abundance [13,21,27].…”