Anti-Phospho-Smad2 (S250) Rabbit Monoclonal Antibody

Scutellarin Inhibits TGF-β1 and Its Downstream Signalling Pathway. a Representative images of Western blotting samples for TGF-β1, p-SMAD2, p-SMAD3, SMAD2/3, SMAD4, p-p38, p38 and p-Erk and Erk1/2 of the mice treated with vehicle, scutellarin or empagliflozin. b – g Quantifications of the protein as indicated. All data are normalized to the STZ group and presented as the mean ± S.D.; n = 4–6 for each group, “n” stands for the number of animals; p vs. the model group (STZ)
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Recombinant HE4 promoted fibroblast-myofibroblast transition and fibroblastic proliferation. A, B Different concentrations of rHE4 facilitated fibronectin, collagen I and α-SMA expression both in mRNA level (n = 3) and protein level (n = 4). The representative western blot images were shown and the band intensity was quantified by ImageJ. C Treatment with rHE4 in HPF cells enhanced phosphorylation of Smad2 (n = 4). D Typical images of EdU-staining HPFs followed by rHE4 treatment for 48 h and the quantitative graph were shown (n = 3). magnification = × 200. E CCK8 proliferation curve signified the pro-proliferative effect of rHE4 (n = 4). HPF was stimulated by rHE4 for 0 h, 24 h, 48 h and 72 h. F Representative western blot images of PCNA and Survivin were displayed and quantified by ImageJ (n = 3). Data are presented as mean ± SEM. P-values were calculated using one-way ANOVA followed by Newman–Keuls test. *P < 0.05, **P < 0.01, and ***P < 0.001 represent significant differences. rHE4, recombinant human epididymis protein 4; EdU, 5-ethynyl-2′-deoxyuridine; CCK8, Cell Counting Kit-8; PCNA, proliferating cell nuclear antigen; HPF, human pulmonary fibroblast
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Western blot analysis of Phospho-Smad2 (S250) using anti-Phospho-Smad2 (S250) antibody (P00090-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human Hela whole cell lysates,
Lane 2: human HepG2 whole cell lysates,
Lane 3: human HT-1080 whole cell lysates,
Lane 4: rat heart tissue lysates,
Lane 5: rat skeletal muscle tissue lysates,
Lane 6: mouse heart tissue lysates,
Lane 7: mouse skeletal muscle tissue lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-Phospho-Smad2 (S250) antigen affinity purified monoclonal antibody (Catalog # P00090-1) at 1:1000 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:1000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for Phospho-Smad2 (S250) at approximately 58 kDa. The expected band size for Phospho-Smad2 (S250) is at 52 kDa.