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. 2018 Feb 15;8:3081. doi: 10.1038/s41598-018-21361-6

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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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SDHB-overexpressing cells deregulates bioenergetic metabolism. (A) ATP assay of SDHB-overexpressing cells. Total extracts were prepared from cells as indicated, and an ATP assay was performed using the ATP Bioluminescence Assay Kit CLSII. (B and C) Δψ_m assay of SDHB-overexpressing cells. Cells were stained with Rhodamine123 and TMRM and analyzed using a flow cytometer. (D,E) ROS assay of SDHB-overexpressing cells. Cells as indicated were treated with CM-H₂DCFDA and DCF and subsequently analyzed using a flow cytometer. (F) Glucose uptake assay of SDHB-overexpressing cells. Cells as indicated were loaded with 2-NBDG and then analyzed using a flow cytometer. (G) LDH activity assay of SDHB-overexpressing cells. Total extracts were prepared from cells and subjected to a LDH activity analysis using a CytoTox 96^® Non-Radioactive Cytotoxicity Assay. (H) Color change and pH measurement of the conditioned media cultured with SDHB-overexpressing cells. Cells as indicated were cultured until confluent and then incubated in fresh media for 24 h. The color and pH of the conditioned media were imaged and measured. The *^, ** and *** represented P-value < 0.05, <0.01 and <0.005, respectively.