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Periodic acceptor excitation spectroscopy of single molecules

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Abstract

Alternating-laser excitation (ALEX) spectroscopy has recently been added to the single-molecule spectroscopy toolkit. ALEX monitors interaction and stoichiometry of biomolecules, reports on biomolecular structure by measuring accurate Förster resonance energy transfer (FRET) efficiencies, and allows sorting of subpopulations on the basis of stoichiometry and FRET. Here, we demonstrate that a simple combination of one continuous-wave donor-excitation laser and one directly modulated acceptor-excitation laser (Periodic Acceptor eXcitation) is sufficient to recapitulate the capabilities of ALEX while minimizing the cost and complexity associated with use of modulation techniques.

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Acknowledgments

We thank N.K. Lee for providing DNA samples and L. LeReste for assistance. This work was funded by NIH grant GM069709-01 to S.W. and A.N.K., DOE grants 02ER63339 and 04ER63938 to S.W., and EPSRC grant EP/D058775, EU Marie Curie Fellowship MIRG-CT-2005-031079, and a UK Bionanotechnology IRC grant to A.N.K. M.H. was supported by a DAAD fellowship.

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Authors and Affiliations

  1. Clarendon Laboratory, Department of Physics and IRC in Bionanotechnology, Oxford University, Oxford, UK

    Sören Doose, Mike Heilemann & Achillefs N. Kapanidis

  2. Department of Chemistry and Biochemistry, University of California Los Angeles, 607 Charles E. Young Drive East, Los Angeles, CA, 90095-1569, USA

    Xavier Michalet & Shimon Weiss

  3. Department of Physiology, University of California Los Angeles, 607 Charles E. Young Drive East, Los Angeles, CA, 90095-1569, USA

    Shimon Weiss

  4. Applied Laser Physics and Laser Spectroscopy, University of Bielefeld, Bielefeld, Germany

    Sören Doose

Authors
  1. Sören Doose
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  2. Mike Heilemann
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  3. Xavier Michalet
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  4. Shimon Weiss
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  5. Achillefs N. Kapanidis
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Correspondence to Sören Doose or Achillefs N. Kapanidis.

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Doose, S., Heilemann, M., Michalet, X. et al. Periodic acceptor excitation spectroscopy of single molecules. Eur Biophys J 36, 669–674 (2007). https://doi.org/10.1007/s00249-007-0133-7

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  • DOI: https://doi.org/10.1007/s00249-007-0133-7

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