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High-Resolution “Fleezers”: Dual-Trap Optical Tweezers Combined with Single-Molecule Fluorescence Detection

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Optical Tweezers

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1486))

Abstract

Recent advances in optical tweezers have greatly expanded their measurement capabilities. A new generation of hybrid instrument that combines nanomechanical manipulation with fluorescence detection—fluorescence optical tweezers, or “fleezers”—is providing a powerful approach to study complex macromolecular dynamics. Here, we describe a combined high-resolution optical trap/confocal fluorescence microscope that can simultaneously detect sub-nanometer displacements, sub-piconewton forces, and single-molecule fluorescence signals. The primary technical challenge to these hybrid instruments is how to combine both measurement modalities without sacrificing the sensitivity of either one. We present general design principles to overcome this challenge and provide detailed, step-by-step instructions to implement them in the construction and alignment of the instrument. Lastly, we present a set of protocols to perform a simple, proof-of-principle experiment that highlights the instrument capabilities.

The original version of this chapter was revised. The grant number in the acknowledgement section has been changed from “RC-105094 (to M.J.C.)” to “MCB-1514706 (to M.J.C.)”

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Acknowledgments

We thank members of the Chemla, Ha, and Comstock laboratories for scientific discussion. Funding was provided by NSF grants MCB-0952442 (CAREER to Y.R.C.), PHY-1430124 (Center for the Physics of Living Cells to Y.R.C.), MCB-1514706 (to M.J.C.), and NIH grant R21 RR025341 (to Y.R.C.).

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Authors and Affiliations

  1. Center for the Physics of Living Cells & Center for Biophysics and Quantitative Biology, University of Illinois, Urbana, IL, USA

    Kevin D. Whitley & Yann R. Chemla

  2. Department of Physics and Astronomy, Michigan State University, East Lansing, MI, USA

    Matthew J. Comstock

Authors
  1. Kevin D. Whitley
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  2. Matthew J. Comstock
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  3. Yann R. Chemla
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Corresponding author

Correspondence to Yann R. Chemla .

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Editors and Affiliations

  1. Department of Anatomy and Structural Biology & Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, Bronx, New York, USA

    Arne Gennerich

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Whitley, K.D., Comstock, M.J., Chemla, Y.R. (2017). High-Resolution “Fleezers”: Dual-Trap Optical Tweezers Combined with Single-Molecule Fluorescence Detection. In: Gennerich, A. (eds) Optical Tweezers. Methods in Molecular Biology, vol 1486. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6421-5_8

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  • DOI: https://doi.org/10.1007/978-1-4939-6421-5_8

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6419-2

  • Online ISBN: 978-1-4939-6421-5

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