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. 2020 Aug 6;10:13297. doi: 10.1038/s41598-020-69990-0

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© The Author(s) 2020

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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Western blots with selected mtDod-PAS-Pep construct ABs. L: Ladder. Lys.: Lysate. OE: protein overexpressing cells. (a) Detection of the target protein in purified form and in the lysate. ABs derived from mtDod-PAS-Pep1 and mtDod-PAS-Pep3 detect target proteins in the lysate. For mtDod-PAS-Pep3, the same blot is shown with two exposure times (13 s and 40 s) to make the potential HSP-110 representing band in the lysate observable. ABs derived from mtDod-PAS-Pep5 and mtDod-PAS-Pep10 did not recognize purified target protein, but seem to recognize a protein in overexpressing cells (uncropped western blots see Supplementary Fig. S11) (b) Comparison with commercially available ABs. Different amounts (1 µg to 60 ng) of purified target protein were loaded and analysed by mtDod-PAS-Pep derived ABs (1 µg/mL) and commercially available ABs (recommended dilutions were used; concentration given when available). All used commercial ABs should recognize different epitopes than the mtDod-PAS-Pep derived ABs, since the given epitope amino acid regions are not overlapping with the selected peptides. Enzo HSP-70: Monoclonal anti HSP70 AB (epitope region amino acids 436–503) from Enzo Life Science, GmbH. *: Concentration assumed based on older aliquots, since information is not given for the newer ones. SCBT HSP-110: Monoclonal anti HSP-105/HSP-110 AB (epitope region amino acids 187–512) from Santa Cruz Biotechnology, Inc. Sigma HSP-4A: Polyclonal anti HSP-4A AB (epitope region amino acids 639–748) from Sigma-Aldrich, Merck KGaA. CellSig. CHIP: Monoclonal anti CHIP AB (epitope region L36 surrounding amino acids (synthetic peptide, length not given)) from Cell Signaling Technology, Inc. For uncropped western blots, see Supplementary Fig. S12. ABs derived from mtDod-PAS-Pep2 also recognize HSP-70. For a comparison of mtDod-PAS-Pep1-3 derived ABs, see Supplementary Fig. S13.