A new Fight-or-Flight Pacemaker Mechanism via Ryanodine Receptor abundance and superclustering
Fig 7
Flowchart of the dSTORM-based RyR cluster analysis algorithm.
Presented is our Single-Molecule Localization Microscopy (SMLM) processing pipeline for precise filtering, delineation, clustering, and statistics of RyR localizations in SANCs. Raw detections are drift-corrected, grouped, and spatially filtered, then gated by localization precision (1–30 nm), photon count (1,000–6,000), PSF half-width (6.42–250 nm), χ² (0–2), and background (≤ mean + 1.5·SD), with a ≥ 6 blinks-per-site rule. Cell region is defined by a periphery mask with nuclear-void removal (if applicable) to compute the final area. The post-processed particles are clustered by DBSCAN using an adaptive nearest-neighbor ε; from the labels we derive nearest neighbor distances (NNDs), cluster-size metrics (mean, SD, 95th percentile), and RyR density, and compare groups by two-sided Mann–Whitney U tests.