Cells were suspended in lysis buffer (0.1% SDS, 40 mM Tris–HCl (pH 7.4), 1% Nonidet P-40, 0.5% sodium deoxycholate, 150 mM NaCl, 4 mM EDTA, 1 mM phenylmethylsulfonyl fluoride, 5 μg/ml aprotinin, 1 μg/ml leupeptin), and were then centrifuged. Lysate samples containing 10 to 20 μg of protein were subjected to SDS-polyacrylamide gel electrophoresis and were then transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, Bedford, MA). PVDF membranes were then incubated overnight at 4°C with primary antibodies and signals were detected using Pierce Western Blotting Substrate (Thermo Scientific, Rockford, IL) according to the manufacturer's instructions. Antibodies against p21, p27, p16, and cyclin dependent kinase (Cdk) 2 were obtained from BD Transduction Laboratories (Franklin Lakes, NJ). Antibodies against FLAG and actin were purchased from Sigma (St Louis, MO). Rb antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA) and Cdk4 antibody was obtained from Epitomics (Burlingame, CA). Phosphorylated-Rb (Ser807/811) antibody was purchased from Cell Signaling Technology (Beverly, MA).
Rb antibody
Manufactured by Santa Cruz Biotechnology
Sourced in United States
The Rb antibody is a laboratory tool used in various research applications. It is designed to specifically recognize and bind to the Rb protein, which plays a crucial role in cell cycle regulation. The Rb antibody can be utilized in techniques such as Western blotting, immunoprecipitation, and immunohistochemistry to detect and study the Rb protein in samples.
Automatically generated - may contain errors
Lab products found in correlation
Pierce western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Polyvinylidene difluoride pvdf membrane, by Merck Group (1 mentions)
Anti parp antibody, by Cell Signaling Technology (1 mentions)
Akt antibody, by Cell Signaling Technology (1 mentions)
Calnexin antibody, by Merck Group (1 mentions)
Pakt ser473 antibody, by Cell Signaling Technology (1 mentions)
P akt antibody, by Cell Signaling Technology (1 mentions)
P extracellular signal regulated kinase erk, by Santa Cruz Biotechnology (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
Sa β gal kit, by Abcam (1 mentions)
Horseradish peroxidase hrp conjugated anti mouse and anti rabbit antibodies, by Merck Group (1 mentions)
Anti erk, by Cell Signaling Technology (1 mentions)
Anti akt, by Cell Signaling Technology (1 mentions)
Anti bcl 2, by Cell Signaling Technology (1 mentions)
Anti caspase 9, by Cell Signaling Technology (1 mentions)
Anti p38, by Cell Signaling Technology (1 mentions)
Ly294002, by Cell Signaling Technology (1 mentions)
Cyclin b1, by Santa Cruz Biotechnology (1 mentions)
Pd98059, by Cell Signaling Technology (1 mentions)
P cdc2, by Santa Cruz Biotechnology (1 mentions)
4 protocols using rb antibody
1
Western Blot Analysis of Cell Cycle Regulators
2
Chalcone-Induced Apoptosis in Liver Cancer
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Poorly differentiated liver cancer cell line Mahlavu cells were cultured in 100 mm culture dish for 24 h. Growth medium was then replaced with IC50 concentrations of chalcones 1 , 9 and 11 or DMSO (control) supplemented medium. cells were incubated for 24 hours then were scraped and collected for western blot analysis. Anti-PARP antibody (Cell Signaling, 9532), pAkt (Ser473) antibody (Cell Signaling, 9271), Akt antibody (Cell Signaling, 9272), pNFkB-p65 (Ser468) antibody (Snat Cruz, sc101750), p21 antibody (Millipore, 05345), pRb (Ser807/811) antibody (Cell Signaling, 9308), Rb antibody (Santa Cruz, sc102), actin antibody (Sigma, A5441) and calnexin antibody (Sigma, C4731) were used as primary antibodies. Anti-rabbit (6154) and anti-mouse (0168) secondary antibodies were used. ImageJ tool (42 (link)) used for protein band intensity comparison. Original Full length blots are given as Supplementary Information.
3
Senescence Regulation by CT Extraction
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CT was obtained from Dr. Park (Kyungnam University, Changwon‐si, Republic of Korea), and CT extraction and separation were performed as previously reported (Shon et al., 2014 ). An SA‐β‐gal kit was purchased from Abcam Inc. (Cambridge, MA, USA). P‐p38, p53, p‐21, p‐silent information regulator 1 (Sirt1), Sirt1, and p‐Akt antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA). Β‐Actin, p‐extracellular signal‐regulated kinase (ERK), and Rb antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX, USA). Horseradish peroxidase (HRP)‐conjugated anti‐mouse and anti‐rabbit antibodies were procured from GeneTex Inc. (Irvine, CA, USA).
4
Investigating Molecular Mechanisms in Cells
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HMH (Fig. 1A ; Sigma-Aldrich Co., St. Louis, MO, USA) was dissolved in 100% dimethyl sulfoxide (DMSO). A 50 mmol/L stock solution of HMH was prepared and stored as small aliquots at −20°C until needed. DMSO, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and horseradish peroxidase (HRP)-conjugated anti-rabbit and anti-mouse antibodies were purchased from Sigma-Aldrich, an apoptosis detection kit was purchased from BD Biosciences (Franklin Lakes, NJ, USA), and 2,7-dichlorofluorescein (DCF) diacetate (H2DCFDA) was purchased from Molecular Probes (Invitrogen, Carlsbad, CA, USA). Phosphospecific anti-JNK, anti-p38, anti-ERK, anti-PI3K, anti-AKT, and anti-mTOR antibodies, anti-caspase-3, anti-caspase-9, anti-poly(ADP-ribose) polymerase (PARP), anti-Bax, anti-Bcl-2, anti-JNK, anti-p38, anti-ERK, anti-PI3K, anti-AKT, and anti-mTOR specific antibodies, the ERK inhibitor PD98059, and the AKT inhibitor LY294002 were purchased from Cell Signaling Technology (Danvers, MA, USA). HRP-conjugated b-actin, p53, p21, p-cdc2, cdc2, cyclin B1, and Rb antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
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