Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using the DC protein assay (Bio-Rad, Hercules, CA). Western blot analysis was performed as described previously [37 (link), 38 (link)]. Antibodies used include Akt1, β-catenin (ser675), N-cadherin, VE-cadherin, Snail1, FoxC2, phospho- and total-Smad 2/3, phospho- and total-p-38 MAPK, GAPDH from Cell Signaling (Danvers, MA)., anti- β-actin, alpha-SMA, and TGFβ2 from Sigma Aldrich (St. Louis, MO) and eNOS from BD Pharmingen (SanDiego, CA). HRP-conjugated goat-anti-mouse and goat-anti-rabbit secondary antibodies were obtained from Bio-Rad (Hercules, CA). Densitometry was done using NIH Image J software.
Phospho and total smad 2 3
Manufactured by Cell Signaling Technology
Phospho- and total-Smad 2/3 is a laboratory equipment product that detects the phosphorylated and total forms of the Smad 2 and Smad 3 proteins. These proteins are key mediators of the transforming growth factor-beta (TGF-β) signaling pathway.
Automatically generated - may contain errors
Lab products found in correlation
Complete lysis buffer, by Merck Group (1 mentions)
Tgf β2, by Merck Group (1 mentions)
Alpha sma, by Merck Group (1 mentions)
Ve cadherin, by Cell Signaling Technology (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
β catenin, by Cell Signaling Technology (1 mentions)
N cadherin, by Cell Signaling Technology (1 mentions)
Snail1, by Cell Signaling Technology (1 mentions)
Anti β actin, by Merck Group (1 mentions)
Dc protein assay, by Bio-Rad (1 mentions)
Protease and phosphatase inhibitor cocktail, by Roche (1 mentions)
Hsp70, by Enzo Life Sciences (1 mentions)
Claudin 5, by Merck Group (1 mentions)
2 protocols using phospho and total smad 2 3
1
Protein Expression Analysis by Western Blot
2
Cardiac Protein and RNA Extraction
Check if the same lab product or an alternative is used in the 5 most similar protocols
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RNA and protein were isolated from the apical portion of the left ventricle using RNeasy (Qiagen) and radioimmunoprecipitation assay buffer with protease inhibitors. The following antibodies were used for immunoblotting: claudin‐5 (#ABT45, Millipore), neuronal nitric oxide synthase (nNOS) (#4234S, Cell Signaling), HSP70 (Enzo Life Sciences), and phospho‐ and total SMAD2/3 (Cell Signaling).
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