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. 2017 Jul 17;7:5582. doi: 10.1038/s41598-017-05817-9

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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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Actin nodules contain PKA signalling proteins. Platelets (2 × 10⁷/ml) were spread on 100 μg/ml fibrinogen for 25 minutes before being washed with PBS. The platelets were then treated with tyrodes with or without 10 nM PGI₂ for a further 10 minutes. The platelets were then fixed, lysed and stained for either (a) PKA RI (1:100), (b) PKA RII (1:100) or (c) pRhoA (1:1000); and co-stained with actin (FITC-Phalloidin) for 60 minutes, before mounting and imaging the slides. Images are representative of at least 3 experiments. Scale bar is 5 μm.