A spatio-temporally compensated acousto-optic scanner for two-photon microscopy providing large field of view

Abstract

Acousto-optic deflectors (AOD) are promising ultrafast scanners for non-linear microscopy. Their use has been limited until now by their small scanning range and by the spatial and temporal dispersions of the laser beam going through the deflectors. We show that the use of AOD of large aperture (13mm) compared to standard deflectors allows accessing much larger field of view while minimizing spatio-temporal distortions. An acousto-optic modulator (AOM) placed at distance of the AOD is used to compensate spatial and temporal dispersions. Fine tuning of the AOM-AOD setup using a frequency-resolved optical gating (GRENOUILLE) allows elimination of pulse front tilt whereas spatial chirp is minimized thanks to the large aperture AOD.

Third-harmonic generation for the study of Caenorhabditis elegans embryogenesis

“…An optimized imaging system that takes into account dispersion for optimizing pulse duration at the sample plane of the microscope, 31 minimizing aberrations 26 ͑such as chromatic͒ and use of objectives optimized for such long wavelengths can be used for extracting more THG signal. If these considerations are taken into account, other scanning configurations could be used to speed up the scanning rates, such as acousto-optical deflectors 32 or a polygonal mirror. 33 To further explore the induced effects of the 1550-nm wavelength, the survival rates of the embryos after long-term imaging exposure were studied.…”

Optical monitoring of neuronal activity at high frame rate with a digital random-access multiphoton (RAMP) microscope

“…However, the use of subpicosecond pulses would be limiting for some applications, like uncaging or photobleaching, that require a high excitation power and small focal volumes. In this case precompensation methods should be used to correct for the dispersion of the laser pulse by AODs (Iyer et al, 2006(Iyer et al, , 2003Kremer et al, 2008;Lechleiter et al, 2002;Reddy and Saggau, 2005;Salomé et al, 2006;Zeng et al, 2006Zeng et al, , 2007.…”

“…Such discontinuous scanning cannot be implemented with galvanometric mirrors but is the essence of AODs, which are digital pointing devices with microsecond switch times (Bullen et al, 1997). Adaptation of AODs to two-photon microscopy has raised important technical difficulties that have been solved in the past years (Iyer et al, 2006(Iyer et al, , 2003Kremer et al, 2008;Lechleiter et al, 2002;Reddy and Saggau, 2005;Salomé et al, 2006;Zeng et al, 2006 leading to the design of functional random-access multiphoton (RAMP) microscopes (Iyer et al, 2006;Salomé et al, 2006;Vucinic and Sejnowski, 2007).…”

Advances in Two-Photon Scanning and Scanless Microscopy Technologies for Functional Neural Circuit Imaging

“…This can be achieved using a single prism [95] or by multiple element systems [91]. AODs, along with some dispersion compensation components, introduce large power losses into the imaging set-up [96]. This can cause the maximum imaging depth to be power limited [91].…”