Miniaturized selective plane illumination microscopy for high-contrast in vivo fluorescence imaging

Abstract

Light-sheet-based fluorescence imaging techniques rely on simultaneous excitation of a single optical plane and thus permit high-contrast optically sectioned imaging of extended tissue samples. Here, we introduce a miniaturized fiber-optic implementation of a selective plane-illumination microscope (miniSPIM). The excitation light was delivered through a single-mode optical fiber, and a light-sheet was created with a cylindrical gradient-index lens and a right-angle microprism. Fluorescence emission was collected orthogonally to the light-sheet through a gradient-index lens assembly and a coherent fiber bundle. The end face of the fiber bundle was imaged onto a charge-coupled device camera. The spatial resolutions of the miniSPIM were 3.2 μm laterally and 5.1 μm axially. Images of fluorescent beads and neurons in mouse neocortex exhibited superior axial resolution and contrast in the miniSPIM-mode compared to images recorded in epi-illumination mode. The miniSPIM may thus enable novel in vivo imaging approaches.