Rats' intestinal mucosa was homogenized on ice-cold water with lysis buffer (Beyotime, P0013 B). Equal protein concentrations were separated by SDS-PAGE (Beyotime, P0050) and transferred to the PVDF membrane (Beyotime, FFP24). After blocking 30 min with the QuickBlock™ Blocking Buffer (Beyotime, P0252), membranes were incubated overnight at 4°C with primary antibodies against MAPK (Abcam, ab185145), IL-1β (Abcam, ab200478), IL-6 (Abcam, ab259341), TGF-β (Cusabio, CSB-PA004279), and TNF-α (Abcam, ab66579). GAPDH (Abcam, ab181602) was used as a normalizing protein. Membranes were then washed 3 times with Western Wash Buffer (Beyotime, P0023C3) 3 times, followed by incubation with horseradish peroxidase (HRP)-conjugated secondary antibody for 2 h. Membranes were then washed 3 times with Western Wash Buffer (Beyotime, P0023C3), then incubated with luminescent substrate solution (Beyotime, P0018M). Protein bands were visualized by Chemiluminescence Imaging System (Bio-Rad, CA, USA) and analyzed with Image J software.
Ab185145
Manufactured by Abcam
Sourced in United Kingdom
Ab185145 is a polyclonal antibody that recognizes the human PARP1 protein. PARP1 is a nuclear enzyme involved in DNA repair and programmed cell death. The antibody can be used for applications such as Western blotting, immunoprecipitation, and immunohistochemistry.
Automatically generated - may contain errors
Lab products found in correlation
Ab200478, by Abcam (1 mentions)
Ab66579, by Abcam (1 mentions)
Quickblock blocking buffer, by Beyotime (1 mentions)
Ffp24, by Beyotime (1 mentions)
Western wash buffer, by Beyotime (1 mentions)
Lysis buffer, by Beyotime (1 mentions)
Chemiluminescence imaging system, by Bio-Rad (1 mentions)
Ab31419, by Abcam (1 mentions)
Ab150075, by Abcam (1 mentions)
Tcs sp8, by Leica (1 mentions)
Ab47363, by Abcam (1 mentions)
Ab181602, by Abcam (1 mentions)
Horseradish peroxidase labeled secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Ab170099, by Abcam (1 mentions)
3 protocols using ab185145
1
Western Blot Analysis of Inflammatory Markers
2
Immunohistochemical Analysis of JNK and p38 in Lung Tissue
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Fresh lung tissue was fixed in 4% paraformaldehyde for 24 h, normally dehydrated and embedded in sections, then the tissue slices were baked at 65 degrees for 60 min, dewaxed and soaked in 4% hydrogen peroxide for 10 min, in 95 degrees citric acid repair solution for 15 min, then closed using goat serum for 1 h, the primary antibody (JNK: ab31419, 1 µg/mL; p38: ab185145, 1/100, abcam, Cambridge, Cambridgeshire, UK) was added dropwise to the tissue slices in proportion, incubated overnight at 4 degrees. Then the corresponding secondary antibody (ab150075, abcam, USA) was added dropwise for 1 h, and finally DAPI staining solution was added dropwise for 15 min, the slices were sealed with anti-burst sealer and photographed by confocal microscopy (LEICA TCS SP8, Weztlar, Germany).
3
Western Blot Analysis of SEMA4C and MAPK
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Lysis buffer was used to extract proteins from HCC tissues or cells. After centrifugation at 12,000 × g, 4°C for 30 min, the supernatant of tissues or cells was measured by BCA kit. Protein specimens (50 µg) were added onto SDS-PAGE and electrophoresed at 60 V. Proteins were then transferred to nitrocellulose filter membranes (NC). Subsequently, the membranes were blocked with skim milk (5-10%) at 37°C for 1 h and then incubated with the primary antibodies against SEMA4C (1:500; sc-136445; Santa Cruz Biotechnology, Inc.), p38 (1:1000, ab170099; Abcam), p-p38 (1:1000, ab47363; Abcam), MAPK (1:1000, ab185145; Abcam), p-MAPK (1:2000, 4370; Cell Signaling Technology, Inc.,) at 4°C overnight. Then, the membranes were incubated with horseradish peroxidase-labeled secondary antibody (1:5000; Santa Cruz Biotechnology) for 1 h at 37°C. GAPDH (1:2000; ab181602; Abcam) was used as the loading control. Finally, enhanced chemiluminescence kit (ECL; EMD Millipore) was used to detect the signals.
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