Single-molecule detection provides researchers with a unique method to probe kinetics of biomolecules in their native environment, without the need to synchronize the molecular states. However, current single-molecule measurements of DNA hybridization kinetics are mostly performed on a surface or inside an electrokinetic trap, which are not physiologically relevant conditions. Recently we demonstrate a time-resolved, 3D single-molecule tracking (3D-SMT) method that that can follow individual DNA molecules diffusing inside a mammalian cell and observe multiple annealing and melting events on the same molecules. By comparing the hybridization kinetics of the same DNA strand in vitro, we found the association constants can be 13- to 163-fold higher in the molecular crowding cellular environment. In contrast to other confocal-feedback 3D single-particle tracking demonstrations, we tracked single DNA reporter strands inside a live cell and measured their annealing-melting kinetics. Although camera-based techniques combined with point-spread function engineering can achieve 3D tracking in live cells, they do not offer any lifetime monitoring capability that can be used to reveal the molecular binding kinetics... ability to design and synthesize nanomaterials with specific photophysical properties is not only a great intellectual challenge, but also one with important practical consequences. To address this challenge, we are currently exploring a new class of biolabels termed few-atom noble metal nanoclusters. Noble metal nanoclusters are collections of small numbers of gold or silver atoms (2-30 atoms) with physical sizes close to the Fermi wavelength of an electron (~0.5 nm for gold and silver). Providing the missing link between atomic and nanoparticle behavior in noble metals, these nanoclusters exhibit optical, electronic, and chemical properties dramatically different from those of much larger nanoparticles or bulk materials. Among those water-soluble noble metal nanoclusters newly developed, DNA-templated silver nanoclusters (DNA/Ag NCs) have attracted great interest in biosensing owing to a number of useful photophysical and photochemical... 

Molecular trafficking within cells, tissues, and engineered 3D multicellular models is critical to the understanding of the development and treatment of various diseases including cancer. However, current tracking methods are either confined to two dimensions or limited to an interrogation depth of ~15 μm. To achieve deep and high-resolution 3D tracking, we have developed a two-photon, 3D single-particle tracking (2P-3D-SPT) method capable of tracking particles at depths up to 200 mm in scattering samples with 22/90 [xy/z] nm spatial localization precision and 50 µs temporal resolution. At shallow depths the localization precision can be as good as 35 nm in all three dimensions. The approach is based on passive pulse splitters used for nonlinear microscopy to achieve spatiotemporally multiplexed 2P excitation and temporally demultiplexed detection to discern the 3D position of the particle.


Recent Publications

  • J. Yang, T. Davis, A.S. Kazerouni, Y.-I. Chen, M.J. Bloom, H.-C. Yeh, T.E. Yankeelov and J. Virostko, "Longitudinal FRET imaging of glucose and lactate dynamics and response to therapy in breast cancer cells", Molecular Imaging and Biology, 2021 .

  • Y.-A. Kuo, C. Jung, Y.-A. Chen, H.-C. Kuo, O.S. Zhao, T.D. Nguyen, J.R. Rybarski, S. Hong, Y.-I. Chen, D.C. Wylie, J.A. Hawkins, J.T. Petty, I.J. Finkelstein and H.-C. Yeh, "Massively parallel selection of NanoCluster Beacons", bioRxiv, 2021. 

  • Y.-I. Chen, Y.-J. Chang, S.-C. Liao, T.D. Nguyen, J. Yang, Y.-A. Kuo, S. Hong, H.-C. Li, Y.-L. Liu, H.G. Rylander III, S.R. Santacruz, T.E. Yankeelov, and H.-C. Yeh, "Deep learning improves complex lifetime estimation in fluorescence lifetime imaging microscopy", Communications Biology, in press.


  • November 24, 2021

    Nina received $300 2022 BPS travel award. Congratulations!

  • November 24, 2021

    Ted received $300 2022 BPS travel award. Congratulations!

  • November 21, 2021

    Ted received $250 Fall 2021 Professional Development Award for attending the 2021 BMES conference. Congratulations!

  • November 17, 2021

    Dr. Yeh gave an invited talk on "Advanced microcopy for cancer research" in the Livestrong Cancer Institutes Basic/translational retreat.

  • November 1, 2021

    Dr. Yeh gave an invited talk on "Using the next-generation sequencing platform for massively parallel selection of fluorescent nanomaterials" in the 2021 SWRM ACS Meeting.


Tim Yeh, Ph.D. (Hsin-Chih Yeh 葉信志)
Associate Professor
Department of Biomedical Engineering
University of Texas at Austin
107 W. Dean Keeton Street Stop C0800
Austin, TX 78712-1801
Office: BME 5.202C
Phone: (512) 471-7931