STED analysis of AZ organization at Drosophila NMJ synapses. (A) Overview of a bouton stained for BRP^N-Term (confocal; magenta) and BRP^Nc82 (STED; green) showing planar (arrow) and vertical (arrowhead) AZs. (B and C) Magnifications of individual planar (left) and vertical (right) AZs stained for BRP^Nc82 (STED) and BRP^N-Term (confocal; B) and BRP^N-Term (STED) and Cac^GFP (confocal; C). (D) Mean normalized planar BRP^N-Term (magenta) and BRP^Nc82 (green) arrangement shown with STED resolution (BRP^N-Term, n = 14; BRP^Nc82, n = 47). (right) The merge superimposed with the intensity profile along one axis through the midpoint for BRP^N-Term (magenta) and BRP^Nc82 (green) is shown. Error bars indicate ± SEM. (E) BRP^Nc82 (STED) and BRP^N-Term (confocal) after expression of full-length BRP cDNA in brp⁶⁹ background (BRP^D1-4). (F and G) Individual planar (left) and vertical (right) AZs stained for BRP^Nc82 (STED) and Cac^GFP (confocal; F) and Cac^GFP (STED) and BRP^Nc82 (confocal; G). All images were deconvolved using Imspector software. Bars: (A) 1 µm; (G) 100 nm.