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Immunoblot analysis

SS Sabato Sorrentino
CI Claudio Iaconetti
SR Salvatore De Rosa
AP Alberto Polimeni
JS Jolanda Sabatino
CG Clarice Gareri
FP Francesco Passafaro
TM Teresa Mancuso
LT Laura Tammè
CM Chiara Mignogna
CC Caterina Camastra
GE Giovanni Esposito
AC Antonio Curcio
DT Daniele Torella
CI Ciro Indolfi
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Protein samples were isolated from HUVEC tranfected with miR-16-Mimic, miR-16-Inhibitor, Mimic-NC or Inhibitor-NC. Protein extracts obtained with NP40 Lysis Buffer (Thermo Fisher Scientific, catalog number #FNN0021) were separated on SDS–PAGE gels and western blot analyses were performed by standard protocols. The antibodies used were the following: polyclonal rabbit anti-eNOS antibody (Cell Signaling, catalog number #9572), goat polyclonal anti-phospho-eNOS (Ser 1177) antibody (Santa Cruz Biotech., sc-12972), AKT antibody (Cell Signaling, catalog number #9272), rabbit Phospho-Akt (Ser473) antibody (Cell Signaling, catalog number #193H12), mouse monoclonal RhoGDIα antibody (Santa Cruz Biotech., sc-373724) and rabbit polyclonal anti-GAPDH (FL-335) antibody (Santa Cruz Biotech., sc-25778). Quantification of Western blots was performed by densitometry using NIH ImageJ software. Full-length blots for the figures were presented in Supplemental Fig. 8.

Copyright and License information: 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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