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Cells were collected in PBS and sequentially lysed in buffer 1 (50 mM HEPES pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% TritonX-100), buffer 2 (10 mM Tris pH 8, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA) and buffer 3 (10 mM Tris pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% sodium deoxycholate, 0.5% N-lauroylsarcosine) with protease inhibitors (Roche)

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PMID: 27358198

ERK1/2 activation in endothelial cells with upregulation of c-Fos, c-Jun, and Egr-1, regulating cell proliferation and vascular tube formation in vitro