Proteins were extracted in lysis buffer according to the manufacturer’s protocol. Lysates were separated by SDS-PAGE and transferred to a nitrocellulose membrane (Bio-Rad, USA). The membranes were incubated in blocking buffer. The membranes were incubated at 4 °C overnight with primary antibodies against the following proteins: AdipoR2 (Abcam, USA), AMPK (Abcam, USA), p-AMPK (Abcam, USA), mTOR (Abcam, USA), p-mTOR (Abcam, USA), S6K (Abcam, USA), pS6K (Abcam, USA), S6P (Abcam, USA)), phosphorylated (Ser240/244) S6 ribosomal protein (pS6P) and GAPDH (CST, USA). Immunoreactivity was visualized with horseradish peroxidase-conjugated goat anti-rabbit antibody (Bioworld, USA). The protein bands were detected and imaged with a ChemiDocXRS+ Gel Imaging System (Bio-Rad, USA) and analysed by densitometric quantification using ImageJ software.
Horseradish peroxidase conjugated goat anti rabbit antibody
Manufactured by Bioworld Technology
Sourced in United States
Horseradish peroxidase-conjugated goat anti-rabbit antibody is a laboratory reagent used in various immunoassays and detection techniques. It is composed of a goat-derived antibody specific to rabbit immunoglobulins, conjugated to the enzyme horseradish peroxidase. This conjugate can be used to detect and quantify the presence of rabbit antibodies in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Chemidoc xrs gel imaging system, by Bio-Rad (3 mentions)
Adipor2, by Abcam (3 mentions)
P ampk, by Abcam (2 mentions)
Nitrocellulose membrane, by Bio-Rad (2 mentions)
P mtor, by Abcam (2 mentions)
P s6k, by Abcam (2 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Ecl reagent, by Bio-Rad (1 mentions)
Radioimmunoprecipitation assay buffer, by Cell Signaling Technology (1 mentions)
Gapdh, by Proteintech (1 mentions)
Anitrocellulose membrane, by Bio-Rad (1 mentions)
4 protocols using horseradish peroxidase conjugated goat anti rabbit antibody
1
Immunoblotting Analysis of Metabolic Signaling Proteins
2
Western Blot Analysis of NUCKS1 Protein
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Western blotting was performed as previously described (31 (link)). Total proteins were isolated by radioimmunoprecipitation assay buffer (Cell Signaling Technologies) and quantified with a bicinchoninic kit (Thermo Fisher Scientific). The isolated proteins were separated by 10% SDS-PAGE prior to semitransfer to polyvinylidene fluoride membranes (Millipore). Next, the membranes were blocked for 1 h in 5% skimmed milk at room temperature and incubated with indicated primary antibodies at 4 °C overnight, followed by incubation with corresponding secondary antibodies for 1.5 h at room temperature. Ultimately, blot bands were visualized via ECL reagent (Bio-Rad), and images were obtained using Tanon 5200-Multi (Tanon Science & Technology Co Ltd).
Antibodies against the following proteins were used for the experiments: NUCKS1 was obtained from Cell Signaling Technology (Danvers) and ABclonal; GAPDH was obtained from Proteintech Group. The secondary antibodies including horseradish peroxidase–conjugated goat anti-rabbit antibody and horseradish peroxidase–conjugated goat antimouse antibody were purchased from Bioworld.
Antibodies against the following proteins were used for the experiments: NUCKS1 was obtained from Cell Signaling Technology (Danvers) and ABclonal; GAPDH was obtained from Proteintech Group. The secondary antibodies including horseradish peroxidase–conjugated goat anti-rabbit antibody and horseradish peroxidase–conjugated goat antimouse antibody were purchased from Bioworld.
3
Protein Expression Analysis in Metabolic Regulation
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Proteins were extracted in lysis buffer according to the manufacturer's protocol. Lysates were separated by SDS-PAGE and transferred to a nitrocellulose membrane (Bio-Rad, USA). The membranes were incubated in blocking buffer.The membranes were incubated at 4°C overnight with primary antibodies against the following proteins:AdipoR2(Abcam,USA),AMPK(Abcam,USA),p-AMPK(Abcam,USA),mTOR(Abcam,USA),p-mTOR(Abcam,USA),S6K(Abcam,USA), pS6K(Abcam,USA), S6P(Abcam,USA)), phosphorylated (Ser240/244) S6 ribosomal protein (pS6P) and GAPDH (CST, USA). Immunoreactivity was visualized with horseradish peroxidase-conjugated goat anti-rabbit antibody (Bioworld, USA). The protein bands were detected and imaged with a ChemiDocXRS + Gel Imaging System (Bio-Rad, USA) and analysed by densitometric quanti cation using ImageJ software.
4
Protein Expression Analysis via Western Blot
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Proteins were extracted inlysis bufferaccording to the manufacturer's protocol. Lysates were separated by SDS-PAGEand transferred to anitrocellulose membrane (Bio-Rad, USA).The membranes were incubated in blocking buffer.Themembranes were incubated with the AdipoR2 (Abcam, USA), AMPK, phosphorylated (Thr172) AMPK (p-AMPK), mTOR, phosphorylated (Ser2448) mTOR (p-mTOR),70-kDa ribosomal protein S6 kinase(S6K), phosphorylated(Thr421/Ser424) p70S6 kinase (pS6K), and S6 ribosomal protein (S6P), phosphorylated(Ser240/244) S6 ribosomal protein (pS6P)and GAPDH (CST, USA) primary antibodyat 4 °Covernight, respectively. Immunoreactivity was visualizedwith horseradish peroxidase-conjugated goat anti-rabbit antibody (Bioworld, USA). Protein bands were detected and imaged with ChemiDocXRS + gel imaging system (Bio-Rad, USA)and analyzed bydensitometric quanti cation using Image J software.
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