Background-free deep imaging by spatial overlap modulation nonlinear optical microscopy

We demonstrate how the resolution and imaging depth limitations of nonlinear optical microscopy can be overcome by modulating the spatial overlap between two-color pulses. We suppress out-of-focus signals, which limit the imaging depth, by a factor of 100, and enhance the lateral and axial resolution by factors of 1.6 and 1.4–1.8 respectively. Using spatial overlap modulation, we demonstrate background-free three-dimensional imaging of fixed mouse brain tissue at depths for which the signals of the conventional technique are swamped by background noise from out-of-focus regions.

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