Proteins for western blot analysis were extracted from the renal cortex by lysing with NETN150 (0.5% NP-40, Tris PH 8.0 50 mM, NaCl 150 mM). The proteins were quantified by Bradford, and the samples were separated by SDS PAGE. Gels were transferred to the nitrocellulose membrane (Axygen, Union City, CA) and then blocked with 5% nonfat milk for 1 hour at room temperature, the membrane was incubated with primary antibodies overnight at 4°C, followed by horseradish peroxidase- (HRP-) linked secondary antibody. The antibodies against TGF-β1 (ab169771) and NF-κBp65 (ab7970) were purchased from Abcam, and phospho-NF-κBp65 (Ser536) was bought from Cell Signaling. After usage of Immobilon Western Chemiluminescent HRP Substrate kit (Millipore Corporation, Billerica, MA), the band was quantified with ImageJ (National Institutes of Health, Bethesda, MD, USA).
Ab169771
Manufactured by Abcam
Sourced in United Kingdom
Ab169771 is a polyclonal antibody that recognizes Human protein HMGB1. It is suitable for use in Western Blot and Immunohistochemistry applications.
Automatically generated - may contain errors
Lab products found in correlation
Ab7970, by Abcam (1 mentions)
Nitrocellulose membrane, by Corning (1 mentions)
Immobilon western chemiluminescent hrp substrate kit, by Merck Group (1 mentions)
Phospho nf κb p65 ser536, by Cell Signaling Technology (1 mentions)
Ab181603, by Abcam (1 mentions)
Nitrocellulose membrane, by Merck Group (1 mentions)
Ab179467, by Abcam (1 mentions)
Ab178695, by Abcam (1 mentions)
Odyssey clx scanner, by LI COR (1 mentions)
2 protocols using ab169771
1
Quantitative Analysis of TGF-β1 and NF-κB in Renal Cortex
2
Protein Quantification and Immunoblotting
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Animal tissues were lysed, and proteins were extracted as described above. After SDS–PAGE, proteins were transferred to nitrocellulose membranes (Millipore, Burlington, MA, USA), which were subsequently blocked with a 5% BSA solution. The membranes were then incubated overnight at 4°C with primary antibodies, including anti-LCP1 (ab236280, 1:1000; Abcam, Cambridge, UK), anti-TGM2 (ab109121, 1:1000; Abcam, UK), anti-TGFBI (ab169771, 1:1000; Abcam, UK), and anti-CSRP2 (ab178695, 1:1000; Abcam, UK). Anti-β-actin (ab179467, 1:5000; Abcam, UK) and anti-GAPDH (ab181603, 1:10000; Abcam, UK) were used as protein loading controls. The specific bands were detected using the LI-COR Odyssey CLx scanner (LI-COR, USA) and further analyzed with ImageJ software.
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