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. 2017 Sep 15;5:72. doi: 10.1186/s40478-017-0474-0

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© The Author(s). 2017

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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Fig. 3 — Correlative immunofluorescence and immunoelectron microscopy to localize HSP27, αB-crystallin and HSP90 in normal and diseased myofibers. (a) HSP27 localization on myofibrils. Top panels: representative immunofluorescence images of CTRL, LGMD2A and MFM-filaminopathy (‘FLN-C’) muscle cells labeled with antibodies to HSP27 (secondary antibody: FITC-conjugated IgG). Samples were counterstained against the PEVK titin (TTN) epitope (secondary antibody: Cy3-conjugated IgG); the merged image is on the right in each group. Bottom panels show nanogold-labeled immunoelectron micrographs. Insets, higher-power images of sarcomeric regions immunostained with anti-HSP27/anti-PEVK. (b) Localization of αB-crystallin (αBC). Top panels: immunofluorescence images of myofibers labeled with anti-αB-crystallin (secondary antibody: Cy3-conjugated IgG) and counterstained for PEVK titin (secondary antibody: FITC-conjugated IgG). Bottom panels show immunoelectron micrographs, insets magnifications. (c) Localization of HSP90. Top panels: immunofluorescence images of myofibers labeled with anti-HSP90 (secondary antibody: Cy3-conjugated IgG) and counterstaining for PEVK titin (secondary antibody: FITC-conjugated IgG). Bottom panels show immunoelectron micrographs, insets magnifications. Bars, 5 μm (confocal images) and 1 μm (EM). For a quantitation of nanogold particle distribution on these and similar immunoelectron micrographs, see Additional file 1: Figure S3