title: Deep and high-resolution three-dimensional tracking of single particles using nonlinear and multiplexed illumination.
abstract: Molecular trafficking within cells, tissues and engineered three-dimensional 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. Here we present a three-dimensional tracking method capable of quantifying rapid molecular transport dynamics in highly scattering environments at depths up to 200 μm. The system has a response time of 1 ms with a temporal resolution down to 50 μs in high signal-to-noise conditions, and a spatial localization precision as good as 35 nm. Built on spatiotemporally multiplexed two-photon excitation, this approach requires only one detector for three-dimensional particle tracking and allows for two-photon, multicolour imaging. Here we demonstrate three-dimensional tracking of epidermal growth factor receptor complexes at a depth of ∼100 μm in tumour spheroids.
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Evan P
authors.initials: EP
authors.lastname: Perillo
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Yen-Liang
authors.initials: YL
authors.lastname: Liu
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Khang
authors.initials: K
authors.lastname: Huynh
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Cong
authors.initials: C
authors.lastname: Liu
authors.affiliation: 1] Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, 1515 Holocombe, Boulevard, Unit 108, Houston, Texas 77030-4009, USA [2] Center for Molecular Medicine and Graduate Institute of Cancer Biology, China Medical University, No. 91 Hsueh-Shih Road, Taichung 40402, Taiwan.
authors.forename: Chao-Kai
authors.initials: CK
authors.lastname: Chou
authors.affiliation: 1] Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, 1515 Holocombe, Boulevard, Unit 108, Houston, Texas 77030-4009, USA [2] Center for Molecular Medicine and Graduate Institute of Cancer Biology, China Medical University, No. 91 Hsueh-Shih Road, Taichung 40402, Taiwan.
authors.forename: Mien-Chie
authors.initials: MC
authors.lastname: Hung
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Hsin-Chih
authors.initials: HC
authors.lastname: Yeh
authors.affiliation: Department of Biomedical Engineering, The University of Texas at Austin, 107 W Dean Keeton Street, C0800, Austin, Texas 78712, USA.
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Dunn
mesh_terms: D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D021621:Imaging, Three-Dimensional; D008958:Models, Molecular; D017711:Nonlinear Dynamics; D061848:Optical Imaging; D021381:Protein Transport
chemical_list: D066246:ErbB Receptors