Proteins were extracted by lysing cells in sodium dodecyl sulfate (SDS) sample buffer (62.5 mM Tris–HCl [pH 6.8], 2% SDS) containing 1 mM phenylmethylsulfonyl fluoride, 10 μg/mL pepstatin, 12.5 μg/mL leupeptin, 2 μg/mL aprotinin, 1 mM sodium orthovanadate, and 1 mM sodium molybdate. Cell extracts were processed for western immunoblotting as described previously [50 (link)]. The following antibodies used for immunoblotting were purchased from the indicated suppliers: mouse monoclonal antibody against AF1q from Abnova (Taipei City, Taiwan), mouse monoclonal antibody against GAPDH from Sigma Aldrich (St Louis, MO), mouse monoclonal antibody against cytokeratin 8/18 from Santa Cruz Biotechnology (Santa Cruz, CA), rabbit polyclonal antibody against Fibronectin from Sigma Aldrich, mouse monoclonal antibody against Vimentin from Abcam (Cambridge, UK), rabbit polyclonal antibody against p38 from Santa Cruz Biotechnology, rabbit polyclonal antibody against Thr180/Tyr182 p38 phosphorylations from Cell Signaling (Danvers, MA), rabbit polyclonal antibody against Erk1/2 from Sigma Aldrich, rabbit polyclonal antibody against Thr202/Tyr204 Erk1/2 phosphorylations from Cell Signaling.
Rabbit polyclonal antibody against fibronectin
Manufactured by Merck Group
Sourced in Macao
Rabbit polyclonal antibody against Fibronectin is a laboratory reagent used to detect and quantify the presence of the extracellular matrix protein fibronectin in biological samples. The antibody is produced by immunizing rabbits with purified fibronectin and collecting the resulting polyclonal antibodies.
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2 protocols using rabbit polyclonal antibody against fibronectin
1
Protein Extraction and Western Blotting
2
Antibodies for Extracellular Matrix Proteins
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A rat monoclonal antibody against mouse EMILIN-1 (clone 1007C11A8)1 (link) was used. Rabbit and guinea pig affinity-purified antibodies for EMILIN-2 and EMILIN-3 were already described6 (link), 20 (link). Affinity purified rabbit polyclonal antibodies directed against mouse fibrillin-1 (pAb9543) and fibrillin-2 (868) were a kind gift from Dr. Lynn Sakai (Shriners Hospital for Children, Portland, OR, USA). Rabbit polyclonal antibody against fibronectin was from Sigma (St Louis, MO). Rabbit polyclonal anti LTBP-1 antibody was a kind gift from Dr. C. H. Heldin (Ludwig Institute for Cancer Research, Uppsala, Sweden). Polyclonal goat anti-LTBP-4 antibody was purchased from R&D Systems. Fibulin-1, -2, -4 and -5 antibodies were generated as previously described50 (link). Recombinant murine C-terminally 2xStrepII tagged full length fibulin-4 and monomeric EMILIN-1 was expressed, and purified as previously described44 (link), 51 (link).
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