Fig. 3: Exchange-PAINT imaging for multicolor and RESI imaging using an SDC-OPR microscope. | Nature Communications

Fig. 3: Exchange-PAINT imaging for multicolor and RESI imaging using an SDC-OPR microscope.

From: Super-resolution imaging in whole cells and tissues via DNA-PAINT on a spinning disk confocal with optical photon reassignment

Fig. 3

a Three-target Exchange-PAINT imaging of microtubules (red), mitochondria (green) and nuclear pore complexes (blue) in U2OS cells, acquired using the SDC-OPR system. Microtubules were imaged via DNA-conjugated anti-alpha-tubulin. NPCs were imaged via Nup96, tagged with mEGFP and labeled with DNA-conjugated anti-GFP nanobodies. Mitochondria were imaged via DNA-conjugated secondary nanobody against TOM20. Sample preparation is described in the methods and imaging conditions are summarized in Supplementary Table 1. Zoomed-in region emphasizes the enhanced resolution obtained on the SDC-OPR system focusing on (1) mitochondria, (2) microtubules and (3) nuclear pore complexes. Scale bars = 1 μm. b Exchange-PAINT image of Nup96-mEGFP proteins stochastically labeled with orthogonal DNA sequences by incubation of the sample with anti-GFP nanobodies, each conjugated with one of four orthogonal sequences. Color represents different rounds of imaging. c Comparison of DNA-PAINT (left) and Resolution Enhancement by Sequential Imaging (RESI, right) for a single NPC illustrating improvement in spatial resolution by RESI. Localizations are rendered as Gaussians with DNA-PAINT localization precision (σDNA-PAINT) and RESI localization precision (σRESI), respectively. A zoomed-in view of a single Nup96 pair, showcasing the enhanced σRESI. Multicolor imaging was repeated in two independent experiments.

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