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European Biophysics Journal

, Volume 36, Issue 6, pp 669–674 | Cite as

Periodic acceptor excitation spectroscopy of single molecules

  • Sören Doose
  • Mike Heilemann
  • Xavier Michalet
  • Shimon Weiss
  • Achillefs N. Kapanidis
Biophysics Letter

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.

Keywords

Single-molecule fluorescence spectroscopy Alternating-laser excitation (ALEX) Förster resonance energy transfer (FRET) Biomolecular interactions 

Notes

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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Copyright information

© EBSA 2007

Authors and Affiliations

  1. 1.Clarendon Laboratory, Department of Physics and IRC in BionanotechnologyOxford UniversityOxfordUK
  2. 2.Department of Chemistry and BiochemistryUniversity of California Los AngelesLos AngelesUSA
  3. 3.Department of PhysiologyUniversity of California Los AngelesLos AngelesUSA
  4. 4.Applied Laser Physics and Laser SpectroscopyUniversity of BielefeldBielefeldGermany

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