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Real-time 3D single molecule tracking

Author

Listed:
  • Shangguo Hou

    (Duke University)

  • Jack Exell

    (Duke University)

  • Kevin Welsher

    (Duke University)

Abstract

To date, single molecule studies have been reliant on tethering or confinement to achieve long duration and high temporal resolution measurements. Here, we present a 3D single-molecule active real-time tracking method (3D-SMART) which is capable of locking on to single fluorophores in solution for minutes at a time with photon limited temporal resolution. As a demonstration, 3D-SMART is applied to actively track single Atto 647 N fluorophores in 90% glycerol solution with an average duration of ~16 s at count rates of ~10 kHz. Active feedback tracking is further applied to single proteins and nucleic acids, directly measuring the diffusion of various lengths (99 to 1385 bp) of single DNA molecules at rates up to 10 µm2/s. In addition, 3D-SMART is able to quantify the occupancy of single Spinach2 RNA aptamers and capture active transcription on single freely diffusing DNA. 3D-SMART represents a critical step towards the untethering of single molecule spectroscopy.

Suggested Citation

  • Shangguo Hou & Jack Exell & Kevin Welsher, 2020. "Real-time 3D single molecule tracking," Nature Communications, Nature, vol. 11(1), pages 1-10, December.
    • Handle: RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-17444-6
    DOI: 10.1038/s41467-020-17444-6
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    Cited by:

    1. Sam Daly & João Ferreira Fernandes & Ezra Bruggeman & Anoushka Handa & Ruby Peters & Sarah Benaissa & Boya Zhang & Joseph S. Beckwith & Edward W. Sanders & Ruth R. Sims & David Klenerman & Simon J. Da, 2024. "High-density volumetric super-resolution microscopy," Nature Communications, Nature, vol. 15(1), pages 1-10, December.
    2. Elias Amselem & Bo Broadwater & Tora Hävermark & Magnus Johansson & Johan Elf, 2023. "Real-time single-molecule 3D tracking in E. coli based on cross-entropy minimization," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    3. Zhuoyang Qin & Zhecheng Wang & Fei Kong & Jia Su & Zhehua Huang & Pengju Zhao & Sanyou Chen & Qi Zhang & Fazhan Shi & Jiangfeng Du, 2023. "In situ electron paramagnetic resonance spectroscopy using single nanodiamond sensors," Nature Communications, Nature, vol. 14(1), pages 1-8, December.
    4. Andrea Bucci & Giorgio Tortarolo & Marcus Oliver Held & Luca Bega & Eleonora Perego & Francesco Castagnetti & Irene Bozzoni & Eli Slenders & Giuseppe Vicidomini, 2024. "4D Single-particle tracking with asynchronous read-out single-photon avalanche diode array detector," Nature Communications, Nature, vol. 15(1), pages 1-12, December.

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