PT - JOURNAL ARTICLE AU - Pauli, Martin AU - Paul, Mila M. AU - Proppert, Sven AU - Sharifi, Marzieh AU - Repp, Felix AU - Kollmannsberger, Philip AU - Sauer, Markus AU - Heckmann, Manfred AU - Sirén, Anna-Leena TI - Targeted volumetric single-molecule localization microscopy of defined presynaptic structures in brain sections AID - 10.1101/568279 DP - 2019 Jan 01 TA - bioRxiv PG - 568279 4099 - http://biorxiv.org/content/early/2019/03/05/568279.short 4100 - http://biorxiv.org/content/early/2019/03/05/568279.full AB - Revealing the molecular organization of anatomically precisely defined brain regions is necessary for the refined understanding of synaptic plasticity. Although, three-dimensional (3D) single-molecule localization microscopy can provide the required molecular resolution, single-molecule imaging more than a few micrometers deep into tissue remains challenging. To quantify presynaptic active zones (AZ) of entire, large, conditional detonator hippocampal mossy fiber (MF) boutons with diameters as large as 10 µm, we developed a method for aberration-free volumetric direct stochastic optical reconstruction microscopy (dSTORM). An optimized protocol for fast repeated axial scanning and efficient sequential labeling of the AZ scaffold Bassoon and membrane bound GFP with Alexa Fluor 647 enables 3D-dSTORM imaging of 25 µm thick mouse brain sections and assignment of AZs to specific neuronal substructures. Quantitative data analysis revealed large differences in Bassoon cluster size and density for distinct hippocampal regions with largest clusters in MF boutons.