Light needle microscopy with spatially transposed detection for axially resolved volumetric imaging

摘要

The demand for rapid three-dimensional volumetric imaging is increasing in various fields, including life science. Laser scanning fluorescence microscopy has been widely employed for this purpose; however, a volumetric image is constructed by two-dimensional image stacking with a varying observation plane, ultimately limiting the acquisition speed. Here we propose a method enabling axially resolved volumetric imaging without a moving observation plane in the framework of laser scanning microscopy. A scanning light needle spot with an extended focal depth provides excitation, which normally produces a deep focus image with a loss of depth information. In our method, the depth information is retrieved from transposed lateral information on an array detector by utilising non-diffracting and self-bending characteristics imposed on fluorescent signals. This technique, implemented in two-photon microscopy, achieves truly volumetric images constructed from a single raster scan of a light needle, which has the capability to significantly reduce the acquisition time.