28 February 2011 Single-wavelength STED microscope
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Abstract
The zero-point STED microscope (US Pat. 5,866,911)1 was the first far-field microscope to overcome the diffraction limit, but optimally it requires two expensive synchronized short-pulsed lasers. Replacing the synchronized pulsed lasers with CW lasers had been proposed to reduce costs1, but this seriously reduced resolution compared to a similarly powered pulsed STED microscope. A recent theoretical and experimental study (Nat. Methods 4, 915 (2007))3 argued that CW STED has better resolution than previously believed, but there appear to be flaws in the theory sufficient to raise questions about its reported experimental confirmation. We describe an alternative approach to reducing cost of the STED microscope while preserving resolution. A portion of the beam from a femtosecond pulsed laser of a wavelength able to excite fluorescence by multiphoton absorption, is passed through a long optical fiber to stretch the pulses to reduce their peak power so they can no longer excite but can quench by stimulated emission. The stretched pulses are shaped into a doughnut profile and then recombined with the first beam for interaction with the specimen. With suitable fluorophores, this instrument should be able to match the resolution performance of the pulsed laser STED microscope using separate lasers. Particularly when added to an existing multiphoton microscope, such performance should be achievable at extremely low added cost.
© (2011) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.
Stephen C. Baer, "Single-wavelength STED microscope", Proc. SPIE 7903, Multiphoton Microscopy in the Biomedical Sciences XI, 79031R (28 February 2011); doi: 10.1117/12.875005; https://doi.org/10.1117/12.875005
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