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. 2018 Mar 29;8:5368. doi: 10.1038/s41598-018-23690-y

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© The Author(s) 2018

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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EXD2 is accessible to added antibody in immunofluorescence without mitochondrial lysis. Immunofluorescent detection following paraformaldehyde fixation requires mitochondrial lysis using for example Triton X100. In the absence of this lysis step mitochondrial matrix proteins and for example mtDNA are not detectable. Thus (panel a) results shows that in the absence of TX100 lysis, EXD2 is detected while neither MRPL12 nor mtDNA are detected by IF. With TX100 lysis, all three are detected. A similar experiment (panel b) shows that both Tomm20 and EXD2, but not mtDNA are detected in the absence of TX100 lysis. A high resolution 20 × 30 µM subsection of a cell using the EXD2 and MRPL12 antibodies illustrates that the EXD2 signal is often enveloping the MRPL12 signal (panel c: some examples are indicated by a white arrow in the merged image), further illustrating EXD2 its outer-membrane localization.