Dewaxing and hydration were performed as we described previously. We used 3% hydrogen peroxide to block endogenous peroxidase activity for 15 min. And the sections were antigen-retrieved in citrate solution by microwave. Following by serum blocking for 1 h and primary antibody including rabbit anti-PER1 (Affinity, USA, 1:100), rabbit anti-MMP13 (Proteintech, China, 1:300) and rabbit anti-NF-κB p65 (Affinity, USA, 1:100) at 4 °C were incubated overnight individually. Secondary antibody was incubated for 1 h after PBS washing. Finally, we used DAB method to visualize the immunoreactive cells and hematoxylin to stain the nucleus. We used ImageJ 1.52i (National Institutes of Health, Germany) to measure the average optical density in anterior, middle and posterior fields of each section.
Rabbit anti mmp13
Manufactured by Proteintech
Sourced in United States
Rabbit anti-MMP13 is a primary antibody that binds specifically to the Matrix Metalloproteinase 13 (MMP13) protein. MMP13 is an enzyme involved in the breakdown of extracellular matrix proteins. This antibody can be used for the detection and quantification of MMP13 in various research applications.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti per1, by Affinity Biosciences (4 mentions)
Rabbit anti nf κb p65, by Affinity Biosciences (2 mentions)
Bca method, by Yeasen (2 mentions)
Rabbit anti nf kb p65, by Affinity Biosciences (2 mentions)
Rabbit anti phospho nf kb p65 ser536, by Affinity Biosciences (2 mentions)
Rabbit anti gapdh, by Affinity Biosciences (2 mentions)
Pvdf membrane, by Merck Group (2 mentions)
Ecl system, by Merck Group (2 mentions)
Anti timp 1, by Proteintech (1 mentions)
6 protocols using rabbit anti mmp13
1
Immunohistochemical Analysis of PER1, MMP13, and NF-κB
2
Quantitative Immunohistochemistry Analysis
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Immunohistochemistry assays were performed using rabbit anti-MMP-13 (dilution 1:200, Proteintech group, Wuhan, China) and anti-TIMP-1 (dilution 1:50, Proteintech group, Wuhan, China). Quantitative analysis was conducted with ImageJ software (version 1.8.0_112, National Institutes of Health, USA).
3
Western Blot Analysis of Protein Expression
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The collected protein samples were quantified using BCA method (Yeasen, China) and separated by 10% SDS-PAGE. Then, the protein was transferred onto the PVDF membrane (Millipore, Germany). After blocking with 5% non-fat milk at room temperature for 1 h, the membrane was incubated with primary antibodies including rabbit anti-PER1 (Affinity, USA, 1:800), rabbit anti-MMP13 (Proteintech, China, 1:1000), rabbit anti-NF-kB p65 (Affinity, USA, 1:500), rabbit anti-Phospho-NF-kB p65 (Ser536) (Affinity, USA, 1:500) and rabbit anti-GAPDH (Affinity, USA, 1:2000) at 4 °C for 16 h. Following the washing with 0.1% TBST, the secondary antibody (Beyotime, China) was incubated with the membrane. After washing with 0.1% TBST, a chemiluminescence ECL system (Millipore, Germany) was used to visualize the immunoreactive proteins.
4
Western Blot Analysis of Protein Expression
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The collected protein samples were quantified using BCA method (Yeasen, China) and separated by 10% SDS-PAGE. Then, the protein was transferred onto the PVDF membrane (Millipore, Germany). After blocking with 5% non-fat milk at room temperature for 1 h, the membrane was incubated with primary antibodies including rabbit anti-PER1 (Affinity, USA, 1:800), rabbit anti-MMP13 (Proteintech, China, 1:1000), rabbit anti-NF-kB p65 (Affinity, USA, 1:500), rabbit anti-Phospho-NF-kB p65 (Ser536) (Affinity, USA, 1:500) and rabbit anti-GAPDH (Affinity, USA, 1:2000) at 4 °C for 16 h. Following the washing with 0.1% TBST, the secondary antibody (Beyotime, China) was incubated with the membrane. After washing with 0.1% TBST, a chemiluminescence ECL system (Millipore, Germany) was used to visualize the immunoreactive proteins.
5
Protein Expression Analysis Protocol
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Cells were lysed using 2% sodium dodecyl sulfate (SDS) lysis buffer (50 mmol/L Tris-HCl, pH 7.4, 2 mmol/L EDTA, 2% SDS) supplemented with protease inhibitor cocktail (Calbiochem, La Jolla, CA, USA) and phenylmethanesulfonyl fluoride (PMSF, Sigma). The protein concentration was determined by the bicinchoninic acid method by using a BCA Protein Assay Kit (Pierce, Rockford, IL, USA). Proteins (50 μg) were fractionated by SDS-polyacrylamide gel electrophoresis and transferred onto a nitrocellulose membrane. The membrane was blocked with 5% nonfat dry milk in TRIS-buffered saline containing 0.1% Tween 20. Rabbit anti-HDAC4 (1:1000 dilution, Cell Signaling Technology, Danvers, MA, USA), mouse anti-MMP3 (1:2000 dilution, ProteinTech), and rabbit anti-MMP13 (1:500 dilution, ProteinTech) antibodies were used to detect the proteins. The blots were developed using a horseradish peroxidase-conjugated secondary antibody and analyzed using enhanced chemiluminescence detection.
6
Immunohistochemical Analysis of PER1, MMP13, and NF-κB
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Dewaxing and hydration were performed as we described previously. We used 3% hydrogen peroxide to block endogenous peroxidase activity for 15 min. And the sections were antigen-retrieved in citrate solution by microwave. Following by serum blocking for 1 h and primary antibody including rabbit anti-PER1 (Affinity, USA, 1:100), rabbit anti-MMP13 (Proteintech, China, 1:300) and rabbit anti-NF-κB p65 (Affinity, USA, 1:100) at 4 °C were incubated overnight individually. Secondary antibody was incubated for 1 h after PBS washing. Finally, we used DAB method to visualize the immunoreactive cells and hematoxylin to stain the nucleus. We used ImageJ 1.52i (National Institutes of Health, Germany) to measure the average optical density in anterior, middle and posterior fields of each section.
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