2D simultaneous spatial and temporal focusing multiphoton microscopy for fast volume imaging with improved sectioning ability

Qiyuan Song, Keisuke Isobe, Kenichi Hirosawa, Katsumi Midorikawa, Fumihiko Kannari

    Research output: Chapter in Book/Report/Conference proceedingConference contribution

    2 Citations (Scopus)

    Abstract

    Simultaneous spatial and temporal focusing (SSTF) multiphoton microscopy offers us widefield imaging with sectioning ability. As extending the idea to 2D SSTF, people can utilize a 2D spectral disperser. In this study, we use a 2D spectral disperser via a virtually-imaged phased-array (VIPA) and a diffraction grating to fulfill the back aperture of objective lens with a spectrum matrix. This offers us an axial resolution enhanced by a factor of ∼1.7 compared with conventional SSTF microscopy. Furthermore, the small free spectral range (FSR) of VIPA will reduce the temporal self-imaging effect around out-of-focus region and thus will reduce the out-of-focus multiphoton excited fluorescence (MPEF) signal of 2D SSTF microscopy. We experimentally show that inside a sample with dense MPEF, the contrast of the sectioning image is increased in our 2D SSTF microscope compared with SSTF microscope. In our microscope, we use a 1 kHz chirped amplification laser, a piezo stage and a sCMOS camera integrated with 2D SSTF to realize high speed volume imaging at a speed of 50 volumes per second as well as improved sectioning ability. Volume imaging of Brownian motions of fluorescent beads as small as 1μm has been demonstrated. Not only the lateral motion but also the axial motion could be traced.

    Original languageEnglish
    Title of host publicationMultiphoton Microscopy in the Biomedical Sciences XV
    EditorsKarsten Konig, Peter T. C. So, Ammasi Periasamy
    PublisherSPIE
    ISBN (Electronic)9781628414196
    DOIs
    Publication statusPublished - 2015 Jan 1
    EventMultiphoton Microscopy in the Biomedical Sciences XV - San Francisco, United States
    Duration: 2015 Feb 82015 Feb 10

    Publication series

    NameProgress in Biomedical Optics and Imaging - Proceedings of SPIE
    Volume9329
    ISSN (Print)1605-7422

    Other

    OtherMultiphoton Microscopy in the Biomedical Sciences XV
    CountryUnited States
    CitySan Francisco
    Period15/2/815/2/10

    Keywords

    • nonlinear microscopy
    • three-dimensional microscopy
    • wide-field microscopy

    ASJC Scopus subject areas

    • Electronic, Optical and Magnetic Materials
    • Biomaterials
    • Atomic and Molecular Physics, and Optics
    • Radiology Nuclear Medicine and imaging

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  • Cite this

    Song, Q., Isobe, K., Hirosawa, K., Midorikawa, K., & Kannari, F. (2015). 2D simultaneous spatial and temporal focusing multiphoton microscopy for fast volume imaging with improved sectioning ability. In K. Konig, P. T. C. So, & A. Periasamy (Eds.), Multiphoton Microscopy in the Biomedical Sciences XV [93292N] (Progress in Biomedical Optics and Imaging - Proceedings of SPIE; Vol. 9329). SPIE. https://doi.org/10.1117/12.2078093