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. 2017 Oct 9;7:12821. doi: 10.1038/s41598-017-13029-4

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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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Expression of iron handling proteins in tubular preparations from 34 week old NZW and NZB/W mice. Proximal and distal nephron tubules were obtained following collagenase digestion and Percoll gradient fractionation of renal cortex. Medullary thick ascending limbs (mTAL) were obtained by collagenase digestion and sieving of outer medullary tissue. Tissue from 3–4 mice were pooled to generate each sample, with data shown as mean ± SEM for n = 8–13 samples per group. In all cases expression of the protein of interest was calculated relative to β-actin and the data normalized to the mean value of the NZW group. (A) Representative Western blot image and group analysis of proximal tubule transferrin receptor (TfR) expression. (B–D) Quantitative real-time PCR analysis of TfR mRNA expression in (B) proximal tubule, (C) mTAL and (D) cortical distal nephron preparations. (E–G) Quantitative real-time PCR analysis of24p3R mRNA expression in (E) proximal tubule, (F) mTAL and (G) cortical distal nephron preparations. (H–J) Representative Western blot images and group analysis of ferroportin (FPN) protein expression in (H) proximal tubule, (I) mTAL and (J) cortical distal nephron preparations. (K–M) Representative Western blot images and group analysis of ferritin protein expression in (K) proximal tubule, (L) mTAL and (M) cortical distal nephron preparations. Groups were compared by Student’s t-test.