Du 897 emccd camera

BMDMs were seeded 24 h before stimulation. Cells were primed with LPS for 4 h, then nigericin was added to activate the NLRP3 inflammasome, and the cells were incubated at 37 °C for 15–45 min. Cells were fixed with 4% paraformaldehyde and processed for STORM imaging. The antibodies used for labelling were anti-DDX3 (Bethyl Laboratories, A300–474A), anti-ASC (Millipore, 04–147) and anti-tubulin (Thermo Fisher Scientific, clone YOL1/34; diluted 1:1,000). STORM acquisition of 10 fields per condition was performed using a Nikon N-STORM system consisting of an inverted TiE stand, a 100 X, 1.45 NA objective, a DU-897 EMCCD camera, an Agilent laser launch and appropriate optics, as previously reported^{42 (link)}. Images of single-molecule reconstructions were exported for downstream analysis by Imaris software. Coordinate (x, y) positions of individual molecules were exported for analysis with a new statistical measure for analysing colocalization in super-resolution images (r-index; X. Liu et al., unpublished protocol). With this method, a value of 1 indicates colocalization, a value of 0 indicates complete randomness and a value of − 1 indicates exclusion.