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. 2017 Jun 6;7:2882. doi: 10.1038/s41598-017-03115-y

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as 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 images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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SOD1^G85R caused neurotoxicity via the formation of insoluble aggregates. (A) Representative fluorescent microscopy images of N2a cells expressing mCherry-SOD1^WT or mCherry-SOD1^G85R. The arrowheads indicate the intracellular SOD1 aggregates. (B) Colocalization of SOD1 and mCherry in the N2a cells. (C) Quantified data of intracellular SOD1 aggregates are expressed as mean ± S.E.M from three independent experiments. In each experiment, at least 200 cells were counted. (D) Immunoblot analysis of SOD1 aggregates. N2a cells expressing SOD1 constructs were lysed with 1% TritonX-100 (Triton soluble fraction; S). Triton-insoluble fraction (I) were resuspended with 2% SDS and then analyzed with immunoblotting by anti-mCherry antibody. (E) N2a cells expressing mCherry-SOD1^WT or mCherry-SOD1^G85R were incubated for 48 h in the differentiation medium. The cell viability was measured by MTT assay. **p < 0.01, ***p < 0.001. Scale bar: 50 µm.