Bio-assay based on single molecule fluorescence detection in microfluidic channels
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A rapid bioassay is described based on the detection of colocalized fluorescent DNA probes bound to DNA targets in a pressure-driven solution flowing through a planar microfluidic channel. By employing total internal reflection excitation of the fluorescent probes and illumination of almost the entire flow channel, single fluorescent molecules can be efficiently detected leading to the rapid analysis of nearly the entire solution flowed through the device. Cross-correlation between images obtained from two spectrally distinct probes is used to determine the target concentration and efficiently reduces the number of false positives. The rapid analysis of DNA targets in the low pM range in less than a minute is demonstrated.
KeywordsBiomedical assay Colocalization Image cross-correlation Microfluidics Single molecule fluorescence TIRF
This work was performed under the auspices of the US Department of Energy by the University of California, Lawrence Livermore National Laboratory under contract number W-7405-Eng-48. This research was funded by the Exploratory Research-Laboratory Directed Research and Development Program at Lawrence Livermore National Laboratory. We would like to thank Jane Bearinger (LLNL) and Stephanie Pasche (ETH Zurich) for graciously providing PLL-g-PEG used in these experiments.
- 19.Ma YF, Shortreed MR, Yeung ES (2000) 72:4640Google Scholar