Instant three-color multiplane fluorescence microscopy

Abstract

One of the most widely used microscopy techniques in biology and medicine is fluorescence microscopy, offering high specificity in labeling as well as maximal sensitivity. For live-cell imaging, the ideal fluorescence microscope should offer high spatial resolution, fast image acquisition, three-dimensional sectioning, and multicolor detection. However, most existing fluorescence microscopes have to compromise between these different requirements. Here, we present a multiplane, multicolor wide-field microscope that uses a dedicated beam splitter for recording volumetric data in eight focal planes and for three emission colors with frame rates of hundreds of volumes per second. We demonstrate the efficiency and performance of our system by three-dimensional imaging of multiply labeled fixed and living cells. The use of commercially available components makes our proposed microscope straightforward for implementation, thus promising for widely used applications.