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. 2019 Jun 19;9:8798. doi: 10.1038/s41598-019-45269-x

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© The Author(s) 2019

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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Tubule calcium-phosphate deposits form in anterior tubules, increase with age and at high culture temperature. Anterior and posterior Malpighian tubule pairs were dissected from adult males (A,C,D), or young adult males and females (B) that had been cultured for 7 or 30 days at 22 °C or 29 °C as indicated on the x-axis. The Pi and calcium (A) or calcium content (B–D) in each tubule pair were measured following tubule dissection and complete lysis of the tissue (mean ± SEM, n = 4–8 tubule pairs, ****p < 0.0001, **p = 0.002).