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STED-SPIM made simple

Teodora Scheul 1 Irène Wang 1 Jean-Claude Vial 1, *
* Corresponding author
1 MOTIV
LIPhy [2011-2015] - Laboratoire Interdisciplinaire de Physique [Saint Martin d’Hères] [2011-2015]
Abstract : We report the development of a stimulated emission depletion (STED) selective plane illumination (SPIM) microscope based on a single diode-pumped solid state (DPSS) laser that simultaneously delivers nanosecond-pulses at two wavelengths. The two wavelengths, 355 nm and 532 nm, are generated by harmonic conversion and they are used to induce respectively excitation and stimulated emission depletion. This source should allow a low-cost, compact, very efficient and simplified STED scheme since the two beams are intrinsically aligned and synchronized. Using a chromatic beam shaping device which leaves the excitation beam unaffected and produces a donut-shaped STED beam, we demonstrate a 300% reduction of the light sheet thickness, together with an enhancement of the sheet uniformity over larger field of view, at low STED power, in Coumarin dye solution. OCIS codes: (180.2520) Fluorescence microscopy; (180.0180) Microscopy. References and links 1. S. W. Hell and J. Wichmann, "Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy," Opt. Lett. 19(11), 780–782 (1994). 2. M. Dyba and S. W. Hell, "Focal spots of size λ/23 open up far-field fluorescence microscopy at 33 nm axial resolution," Phys., "High-resolution three-dimensional imaging of large specimens with light sheet-based microscopy," Nat. Methods 4(4), 311–313 (2007). 8.
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Teodora Scheul, Irène Wang, Jean-Claude Vial. STED-SPIM made simple. Optics Express, Optical Society of America - OSA Publishing, 2014, 88, pp.163901 - 915. ⟨10.1364/OE.22.030852⟩. ⟨hal-01091510⟩

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