Western blotting was performed as described previously [22 (link)]. Briefly, whole cell lysates were electrophoresed using 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) gels and were transferred to a polyvinylidene difluoride (PVDF) membrane. The membrane was then incubated with primary antibodies against P21 antibody (1:2000; Cell Signaling Technology, Danvers, Massachusetts, USA), cyclin D1 antibody (1:2000; Cell Signaling Technology, Danvers, Massachusetts, USA), cyclin E1 (1:2000; Cell Signaling Technology, Danvers, Massachusetts, USA), CDK4 antibody (1:1000; Cell Signaling Technology, Danvers, Massachusetts, USA), CDK6 antibody (1:1000; Cell Signaling Technology, Danvers, Massachusetts, USA), KI67 antibody (1:3000; Servicebio, Wuhan, Hubei, China) and EZH2 antibody (1:2000; Cell Signaling Technology, Danvers, Massachusetts, USA). The membranes were then incubated with the corresponding secondary antibodies (Cell Signaling Technology, Boston, MA, USA) and visualized using an enhanced chemiluminescence kit (Vazyme, Nanjing, Jiangsu, China).
P21 antibody
Manufactured by Cell Signaling Technology
Sourced in United States
The P21 antibody is a tool used in research laboratories to detect the presence and quantify the levels of the P21 protein. P21 is a cyclin-dependent kinase inhibitor that plays a crucial role in cell cycle regulation. The antibody can be used in various immunoassay techniques, such as Western blotting and immunohistochemistry, to study the expression and localization of P21 in different biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence kit, by Vazyme (1 mentions)
Ezh2 antibody, by Cell Signaling Technology (1 mentions)
Cyclin d1 antibody, by Cell Signaling Technology (1 mentions)
Cdk6 antibody, by Cell Signaling Technology (1 mentions)
Ki 67 antibody, by Wuhan Servicebio Technology (1 mentions)
Cyclin e1, by Cell Signaling Technology (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Gapdh, by Boster Bio (1 mentions)
Vimentin, by Boster Bio (1 mentions)
C myc, by Affinity Biosciences (1 mentions)
Cyclin d1, by Affinity Biosciences (1 mentions)
β catenin, by Affinity Biosciences (1 mentions)
N cadherin, by Affinity Biosciences (1 mentions)
β actin, by Boster Bio (1 mentions)
E cadherin antibody, by BD (1 mentions)
Sds polyacrylamide gel, by Bio-Rad (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Bio-Rad (1 mentions)
Supersignal west pico chemiluminescent substrate detection reagent, by Thermo Fisher Scientific (1 mentions)
P53 antibody, by Abcam (1 mentions)
P16 antibody, by Cell Signaling Technology (1 mentions)
6 protocols using p21 antibody
1
Western Blot Analysis of Cell Cycle Regulators
2
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total tissues and cellular proteins were extracted and separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred onto polyvinylidene fluoride membranes (Millipore, USA). The membranes incubated overnight at 4 °C with primary antibodies. The antibodies against CDK2, Cyclin D1, β-catenin, C-myc, N-cadherin, Snail were purchased from Affinity (USA), p21 antibody was from Cell Signaling Technology (USA), E-cadherin antibody was from BD Biosciences (USA), antibodies against MMP9, Vimentin, β-actin and GAPDH were from Boster (China).
3
Western Blot Analysis of p21 and TET2
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell pellets were lysed in 1 × sample buffer containing phosphatase and protease inhibitors. The lysates were sonicated, quantified and boiled for 10 min. Equal amount was electrophoresed in 7.5% or 4–20% SDS–polyacrylamide gels (Bio-Rad, Hercules, CA, USA). After transferring the proteins on to nitrocellulose membrane, filters were incubated in 5% non-fat dry milk in PBST (1X PBS plus 0.2% Tween-20) for 1 h. Blot was probed with p21 antibody (1:1000 dilution, Cell Signaling Technology, Danvers, MA) or TET2 antibody (EMD Millipore) in 1:2000 dilution and ß-actin antibody (1: 10,000 dilution) overnight at 4 °C in PBST, and subsequently incubated with a horseradish peroxidase-conjugated secondary antibody (BioRad) for 1 h at room temperature. Immuno-bound antibodies were detected by Super Signal West Pico Chemiluminescent Substrate detection reagent (Pierce/Thermo Scientific, Rockford, IL, USA) and visualized by autoradiography. The band density on the scanned images was quantified by ImageJ software (ImageJ.net).
4
Senescence Regulation in Metabolic Disorders
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following drugs and reagents were used in the study: DMEM/F12 media without phenol red (Gibco), fetal bovine serum (FBS, Sciencell), RNA isolation kit, Color SYBR Green qPCR Master Mix and 4× reverse transcription Mix (EZBioscience, A0012), enzyme linked immunosorbent assay (ELISA) kits (E2, Labor Diagnostika Nord, FR E-2000; FSH, Immunoway, KE1425; LH, Immunoway, KE1475; AMH, Jingmei, JM11692M1; insulin, Alpco, 80-INSMSU-E01, E10), Normal rabbit IgG (CST, 2729), Normal mouse IgG (CST, 7076), p53 antibody (Abcam ab131442), p16 antibody (Cell Signaling Technology Cat, sc1661), p21 antibody (Cell Signaling Technology, sc-6246), AGER antibody (Abcam, ab3611), Senescenceβ-Galactosidase Staining Kit (Beyotime, C0602), Cell Cycle Staining Kit (Multi Sciences, CCS021), insulin (Absin, abs42019847), metformin (Topscience, T8526), high-fat diet (Jiangsu Xietong Pharmaceutical Bio-engineering Co., Ltd., D12451).
5
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were harvested and resuspended in cell lysis buffer (150 mM NaCl, 50 mM HEPES (pH 7.5), 1% NP40) containing a protease inhibitor cocktail (Roche, Basel, Switzerland). Whole-cell lysates were resolved by SDS-PAGE and transferred to PVDF membranes (GE Healthcare, Uppsala, Sweden) via Western blotting. Proteins were detected with a 1:1000 or 1:5000 dilution of the primary antibody using a chemiluminescence system (Dogen, Seoul, Korea). Images were acquired using the LAS4000 system (GE Healthcare, Uppsala, Sweden). The CTRP1 antibody was purchased from Invitrogen (Carlsbad, CA, USA), p21 antibody from Cell Signaling Technology (Danvers, MA, USA), and p53 antibody from Santa Cruz Biotechnology (Dallas, TX, USA). FR180204, ERK inhibitor, was purchased from Calbiochem (San Diego, CA, USA).
6
Western Blotting Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting analysis was as described in our previous article.22 (link) Briefly, 72 hr after transfection, cells were lysed with radioimmunoprecipitation assay (RIPA) buffer (Sigma-Aldrich, St. Louis, MO, USA), and then western blotting was carried out with standard procedures. The primary antibodies included SETDB1 antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), p53 antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), p21 antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), PUMA antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), Gadd45 antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), γ-H2AX antibody (1:1,000; Cell Signaling Technology, Boston, MA, USA), and GAPDH antibody (1:2,000; Santa Cruz Biotechnology). The secondary antibodies included anti-mouse horseradish peroxidase (HRP)-conjugated antibody (1:5,000; Bio-Rad, Hercules, CA, USA) or anti-rabbit-HRP (1:5,000; Cell Signaling Technology, Boston, MA, USA).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!