Abstract
Fast volumetric sampling of biological specimens with laser scanning microscopes is required in many fields of neurobiology and medicine. When image acquisition in the lateral (x,y) plane reaches frame rates up to 100Hz by means of resonant scanner systems, a jump between two z-layers in the image stack should also be performed in a few milliseconds, a time scale that cannot be reached by moving the objective or the specimen. Several proposed remote focusing systems only translate a small mirror with a mass below 1g and therefore rely on fast linear positioners. We present measurements with a fast remote focusing system on a two-photon-microscope using a voice coil driven linear stage that we developed. The voice coil stage has a travel range of 1000μm, a 10μm step response time of 1ms and a position accuracy better than 1 μm. We characterize the dynamic properties of the voice coil stage and describe the remote focus system. As proof of concept, we show its applicability for 2-photon calcium imaging of neuronal activity in the CA1 region of rat hippocampus. The remote focusing system is compatible with the open-source software ScanImage (HHMI Janelia Research Campus/Vidrio Technologies, LCC).
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