Journal article
Precision and accuracy of single-molecule FRET measurements—a multi-laboratory benchmark study
- Abstract:
- Single-molecule Förster resonance energy transfer (smFRET) is increasingly being used to determine distances, structures, and dynamics of biomolecules in vitro and in vivo. However, generalized protocols and FRET standards to ensure the reproducibility and accuracy of measurements of FRET efficiencies are currently lacking. Here we report the results of a comparative blind study in which 20 labs determined the FRET efficiencies (E) of several dye-labeled DNA duplexes. Using a unified, straightforward method, we obtained FRET efficiencies with s.d. between ±0.02 and ±0.05. We suggest experimental and computational procedures for converting FRET efficiencies into accurate distances, and discuss potential uncertainties in the experiment and the modeling. Our quantitative assessment of the reproducibility of intensity-based smFRET measurements and a unified correction procedure represents an important step toward the validation of distance networks, with the ultimate aim of achieving reliable structural models of biomolecular systems by smFRET-based hybrid methods.
- Publication status:
- Published
- Peer review status:
- Peer reviewed
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(Preview, Version of record, pdf, 497.6KB, Terms of use)
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(Preview, Version of record, pdf, 4.1MB, Terms of use)
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- Publisher copy:
- 10.1038/s41592-018-0085-0
Authors
- Publisher:
- Springer Nature Publishing Group
- Journal:
- Nature Methods More from this journal
- Volume:
- 15
- Pages:
- 669-676
- Publication date:
- 2018-08-31
- Acceptance date:
- 2018-05-25
- DOI:
- EISSN:
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1548-7105
- ISSN:
-
1548-7091
- Pmid:
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30171252
- Language:
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English
- Keywords:
- Pubs id:
-
pubs:912075
- UUID:
-
uuid:4e671113-967a-4fbd-bb06-dfcf381cfd8e
- Local pid:
-
pubs:912075
- Source identifiers:
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912075
- Deposit date:
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2019-02-06
Terms of use
- Copyright holder:
- Hellenkamp et al
- Copyright date:
- 2018
- Notes:
-
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.
Note: an erratum exists for this article, originally published and available at: https://doi.org/10.1038/s41592-018-0193-x
- Licence:
- CC Attribution (CC BY)
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