Fig. 2: Implementation of C-MELM using a digital micromirror device (DMD).

a Experimental setup consisting of a commercial wide-field epi-fluorescence microscope with a modified excitation path (Methods): The excitation light (green) is reflected off the DMD before it is relayed by a telescope, undergoes diffraction through the objective and results in structured illumination in the sample plane. See Supplementary Fig. 2 for details. b Subset of the DMD array configured in a periodic series of “on” and “off”-mirrors. c, d Example characterization of the structured illumination using a single 46-nm fluorescent bead (Methods). c We translated a fluorescent bead relatively to the structured illumination generated with the DMD configuration in b and recorded the number of photons in its resulting diffraction-limited spots (top row) for each position (blue dots) during 50-ms exposures. For each position of the bead, this was repeated for a total of three phases of the structured illumination (colored dots). A harmonic function with parameters obtained by a fit to data (colored curves, Methods, Supplementary Note 3) describes the data perfectly and thus characterizes the structured illumination. For each position, the total intensities were normalized by the total photon number observed in the three diffraction-limited spots recorded at that position. Error bars represent standard error of the calculated relative photon number. d Experimental bead images (top row) obtained at a single stage position (gray area in c) under illumination from three different phases as in c. All three images are fitted with a point-spread function model (bottom row) for a point-source under assumption that its position is identical in the three images (Methods, Supplementary Note 3). This enables accurate estimation of total intensity also in dim images (e.g., for Phase 2). The effective pixel size was 107 nm.