Wide-field depth-sectioning fluorescence microscopy using projector-generated patterned illumination

摘要

We present a simple and cost-effective wide-field, depth-sectioning, fluorescence microscope utilizing a commercial multimedia projector to generate excitation patterns on the sample. Highly resolved optical sections of fluorescent pollen grains at 1.9 μm axial resolution are constructed using the structured illumination technique. This requires grid excitation patterns to be scanned across the sample, which is straightforwardly implemented by creating slideshows of gratings at different phases, projecting them onto the sample, and synchronizing camera acquisition with slide transition. In addition to rapid dynamic pattern generation, the projector provides high illumination power and spectral excitation selectivity. We exploit these properties by imaging mouse neural cells in cultures multistained with Alexa 488 and Cy3. The spectral and structural neural information is effectively resolved in three dimensions. The flexibility and commercial availability of this light source is envisioned to open multidimensional imaging to a broader user base.