The following were purchased from Sigma-Aldrich (Dorset, UK): Dulbecco’s Modified Eagle Medium (DMEM), Penicillin/Streptomycin, MEM non-essential amino acids 100×, 0.25% Trypsin-EDTA, L-glutamine, putrescine, spermidine, spermine. Fetal bovine serum (FBS) was from Gibco by Life Technologies (Sao Paolo, Brazil), mTOR siRNA and negative control were from Fisher Chemical (Loughborough, UK), protease and phosphatase inhibitors cocktail from Roche (Indianapolis, IN, USA), BCA protein assay kit from BioVision (Milpitas, CA, USA). RIPA lysis buffer 10× from Millipore (Burlinton, MA, USA). The following antibodies were purchased from Abcam Biotechnology (Cambridge, UK) and used at the indicated dilutions: rabbit polyclonal anti-actin antibody (ab119716, 1:7, 500 dilution), goat polyclonal anti-rabbit IgG-HRP (ab205718, 1:10,000 dilution). Additionally, mouse monoclonal anti-ODC antibody (sc-398116, 1:100 dilution), mouse monoclonal anti-phospho-4EBP1 (sc-293124, 1:200 dilution), mouse monoclonal anti-phospo-p70S6K (sc-8416, 1:300 dilution), mouse monoclonal anti- eIF4E (sc-271480, 1:300 dilution) and goat anti-mouse IgG-HRP (sc-2005, 1:2,500 dilution) antibodies were from Santa-Cruz Biotechnology (Heidelberg, Germany). Rapamycin and the class 1 dual inhibitors of PI3K and mTORC1, NVP-BEZ235, were from Cayman Chemicals (Ann Arbor, MI, USA).
Goat polyclonal anti rabbit igg hrp
Manufactured by Abcam
Sourced in United Kingdom
Goat polyclonal anti-rabbit IgG-HRP is a secondary antibody conjugated with horseradish peroxidase (HRP). It is designed to detect and bind to rabbit primary antibodies in immunoassays and immunoblotting applications.
Automatically generated - may contain errors
Lab products found in correlation
Protease and phosphatase inhibitors cocktail, by Roche (2 mentions)
Ripa lysis buffer 10, by Merck Group (2 mentions)
Goat anti mouse igg hrp sc 2005, by Santa Cruz Biotechnology (2 mentions)
Dulbecco s modified eagle medium dmem, by Merck Group (2 mentions)
Penicillin streptomycin, by Merck Group (2 mentions)
L glutamine, by Merck Group (2 mentions)
Rapamycin, by Cayman Chemical (1 mentions)
Bca protein assay kit, by Abcam (1 mentions)
Rabbit anti actin polyclonal antibody, by Abcam (1 mentions)
Nvp bez235, by Cayman Chemical (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Chemiluminescence detection kit, by Merck Group (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Sds polyacrylamide gel, by Beyotime (1 mentions)
Mmp 9, by Cell Signaling Technology (1 mentions)
P erk, by Abcam (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Spermidine, by Merck Group (1 mentions)
Putrescine, by Merck Group (1 mentions)
Foetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
3 protocols using goat polyclonal anti rabbit igg hrp
1
Regulation of mTOR Signaling Pathway
2
Western Blot Analysis of Apoptosis and Signaling
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Cells lysates were prepared with radioimmunoprecipitation assay buffer (RIPA) lysis buffer supplement with protease inhibitor cocktail (Calbiochem) and phosphatase inhibitor (Roche). The proteins are electrophoresed on 6–15% SDS-polyacrylamide gel (Beyotime Biotechnology) and transferred onto polyvinylidene fluoride membranes (Millipore, Billerica, MA, USA). The membranes were blocked in the TBS solution (100 mM Tris, 150 mM NaCl, pH 7.6) containing 5% (w/v) skim milk, and 0.5% (v/v) Tween-20 for 1 hour at room temperature. The membranes were incubated with primary antibodies overnight at 4°C followed by incubation with secondary antibodies for 1 hour. Primary antibodies used in this study were antibodies against caspase-3, -8, -9, Bax, pEGFR, pVEGF-R2, pAKT, AKT, and NK-κB, metal matrix protease (MMP) 2, MMP9, β-actin (Cell Signaling Technology, USA), pERK, and ERK (Abcam, UK). Goat polyclonal anti-mouse IgG-horseradish peroxidase (HRP) or goat polyclonal anti-rabbit IgG-HRP were used as secondary antibodies (Abcam, UK). Bands were visualized by chemiluminescence detection kit (Millipore, USA). All experiments were repeated 3 times.
3
Polyamine Signaling Pathway Analysis
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The following were purchased from Sigma-Aldrich (Dorset, UK): Dulbecco’s Modified Eagle Medium (DMEM), Penicillin/Streptomycin, MEM non-essential amino acids 100×, Trypsin-EDTA, L-glutamine, putrescine, spermidine, and spermine. Foetal bovine serum (FBS) was from Gibco Life Technologies, Sao Paolo, Brazil. The protease and phosphatase inhibitors cocktail were from Roche, Indianapolis, IN, USA. RIPA lysis buffer 10× from Millipore, Burlington, VT, USA. The following antibodies were purchased from Abcam Biotechnology, Cambridge, UK: rabbit polyclonal anti-β-actin (ab119716, 1:7500 dilution), goat polyclonal anti-rabbit IgG-HRP (ab205718, 1:10,000 dilution), goat polyclonal anti-AZ1 (ab223481, 1:300), donkey anti-goat IgG H&L-HRP (ab97110, 1:2000). Mouse monoclonal anti-ODC (sc-398116, 1:100 dilution) and goat anti-mouse IgG-HRP (sc-2005, 1:2500 dilution) antibodies were from Santa-Cruz Biotechnology, Heidelberg, Germany.
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