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    The power of single and multibeam two-photon microscopy for high-resolution and high-speed deep tissue and intravital imaging.

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    Authors
    Niesner, Raluca
    Andresen, Volker
    Neumann, Jens
    Spiecker, Heinrich
    Gunzer, Matthias
    Issue Date
    2007-10-01
    
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    Abstract
    Two-photon microscopy is indispensable for deep tissue and intravital imaging. However, current technology based on single-beam point scanning has reached sensitivity and speed limits because higher performance requires higher laser power leading to sample degradation. We utilize a multifocal scanhead splitting a laser beam into a line of 64 foci, allowing sample illumination in real time at full laser power. This technology requires charge-coupled device field detection in contrast to conventional detection by photomultipliers. A comparison of the optical performance of both setups shows functional equivalence in every measurable parameter down to penetration depths of 200 microm, where most actual experiments are executed. The advantage of photomultiplier detection materializes at imaging depths >300 microm because of their better signal/noise ratio, whereas only charge-coupled devices allow real-time detection of rapid processes (here blood flow). We also find that the point-spread function of both devices strongly depends on tissue constitution and penetration depth. However, employment of a depth-corrected point-spread function allows three-dimensional deconvolution of deep-tissue data up to an image quality resembling surface detection.
    Citation
    The power of single and multibeam two-photon microscopy for high-resolution and high-speed deep tissue and intravital imaging. 2007, 93 (7):2519-29 Biophys. J.
    Affiliation
    Helmholtz Centre for Infection Research, Junior Research Group Immunodynamics, D-38124 Braunschweig, Germany.
    Journal
    Biophysical journal
    URI
    http://hdl.handle.net/10033/16233
    DOI
    10.1529/biophysj.106.102459
    PubMed ID
    17557785
    Type
    Article
    Language
    en
    ISSN
    0006-3495
    ae974a485f413a2113503eed53cd6c53
    10.1529/biophysj.106.102459
    Scopus Count
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