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Nanoprobes for Live-Cell Gene Detection

Chapter

Abstract

The ability to visualize in real time the expression dynamics and localization of specific RNAs in vivo offers tremendous opportunities for biological and disease studies including cancer detection. However, quantitative methods such as real-time polymerase chain reaction (PCR) and DNA microarrays rely on the use of cell lysates thus not able to obtain important spatial and temporal information. Fluorescence proteins and other reporter systems cannot image endogenous RNA in living cells. Fluorescence in situ hybridization (FISH) assays require washing to achieve specificity, therefore can only be used with fixed cells. Here we review the recent development of nanostructured probes for living cell RNA detection and discuss the biological and engineering issues and challenges of quantifying gene expression in vivo. In particular, we describe methods that use oligonucleotide probes, combined with novel delivery strategies, to image the relative level, localization, and dynamics of RNA in live cells. Examples of detecting endogenous messenger RNAs, as well as imaging their subcellular localization are given to illustrate the biological applications, and issues in probe design, delivery, and target accessibility are discussed. The nanostructured probes promise to open new and exciting opportunities in sensitive gene detection for a wide range of biological and medical applications.

Keywords

Hairpin probe Oligonucleotide probe RNA detection Live cell Molecular beacon Fluorescence resonance energy transfer 

Notes

Acknowledgments

This work was supported by the National Heart Lung and Blood Institute of the NIH as a Program of Excellence in Nanotechnology (HL80711), by the National Cancer Institute of the NIH as a Center of Cancer Nanotechnology Excellence (CA119338), and by the NIH Roadmap Initiative in Nanomedicine through a Nanomedicine Development Center award (PN2EY018244).

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© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Department of Biomedical EngineeringGeorgia Institute of Technology and Emory UniversityAtlantaUSA
  2. 2.Department of Biomedical EngineeringGeorgia Institute of TechnologyAtlantaUSA
  3. 3.Department of BioengineeringUniversity of PennsylvaniaPhiladelphiaUSA

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