Cell lysates (30 μg protein each sample) dissolved in Laemmli buffer 5×, boiled for 5 min were resolved in 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis gels (SDS-PAGE, Bio-Rad Laboratories, Hercules, CA, USA). After electrophoresis they were transferred to polyvinylidene fluoride membranes (PVDF, Bio-Rad Laboratories), which were incubated overnight at 4 °C with specific primary antibodies: anti phospho-Akt (p-Akt; 1:1000; Ser473, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), anti Akt (1:1000; Santa Cruz Biotechnology), anti UCP1 (1:1000; 4E5, Santa Cruz Biotechnology), anti p-AMPKα/β (1:1000; D-6, Santa Cruz Biotechnology), AMPKα/β (1:1000; Santa Cruz Biotechnology), anti Mfn2 (1:1000; H-68, Santa Cruz Biotechnology). The membranes were washed and then incubated with horseradish peroxidase-coupled goat anti-rabbit IgG (Sigma), peroxidase-coupled rabbit anti-goat IgG and horseradish peroxidase-coupled goat anti-mouse IgG (Sigma) for 45 min and were developed through a nonradioactive method using Western Lightning Chemiluminescence (PerkinElmer Life and Analytical Sciences, Waltham, MA, USA). Phosphorylated protein expression was calculated as a ratio towards specific total protein or β-actin (1:5000; C4, Santa Cruz Biotechnology).
Horseradish peroxidase coupled goat anti mouse igg
Manufactured by Merck Group
Sourced in United States
Horseradish peroxidase-coupled goat anti-mouse IgG is a laboratory reagent used to detect the presence of mouse immunoglobulin G (IgG) in various biological samples. It consists of goat-derived antibodies that specifically bind to mouse IgG, coupled with the enzyme horseradish peroxidase. This enzyme can be used to catalyze a colorimetric or luminescent reaction, allowing for the visualization and quantification of the target mouse IgG.
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Lab products found in correlation
Peroxidase coupled goat anti rabbit igg, by Merck Group (2 mentions)
Horseradish peroxidase coupled goat anti rabbit igg, by Merck Group (2 mentions)
Anti akt, by Santa Cruz Biotechnology (2 mentions)
Sds page, by Bio-Rad (1 mentions)
Polyvinylidene fluoride (pvdf), by Bio-Rad (1 mentions)
Polyvinylidene fluoride membrane, by Bio-Rad (1 mentions)
Ser1177, by Cell Signaling Technology (1 mentions)
Anti enos, by Cell Signaling Technology (1 mentions)
Anti erk1 2, by Santa Cruz Biotechnology (1 mentions)
Anti inos, by Santa Cruz Biotechnology (1 mentions)
Anti cleaved caspase 9, by Abcam (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
Anti cytochrome c, by Santa Cruz Biotechnology (1 mentions)
Anti phospho akt, by Santa Cruz Biotechnology (1 mentions)
Anti phospho erk1 2, by Santa Cruz Biotechnology (1 mentions)
Anti phospho enos, by Cell Signaling Technology (1 mentions)
2 protocols using horseradish peroxidase coupled goat anti mouse igg
1
Western Blot Analysis of Signaling Proteins
2
Western Blot Analysis of Signaling Proteins
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Cell lysates (30 μg of each protein sample) were loaded onto sodium dodecyl sulfate polyacrylamide gel electrophoresis and, thereafter, they were transferred to polyvinylidene fluoride membranes (Bio-Rad Laboratories), which were incubated with specific primary antibodies: anti phospho-eNOS (1:1000; Ser1177, Cell Signaling Technologies), anti eNOS (1:1000; Cell Signaling Technologies), anti iNOS (1:500; Santa Cruz Biotechnology), anti phospho-Akt (1:1000; Ser 473, Santa Cruz Biotechnology), anti Akt (1:1000; Santa Cruz Biotechnology) anti phospho-ERK 1/2 (1:1000; Thr 202/Tyr 204, Santa Cruz Biotechnology), anti ERK 1/2 (1:1000; Santa Cruz Biotechnology), anti Cleaved Caspase-9 (1:1000; Abcam, Cambridge, UK) and anti Cytochrome C (1:1000, Santa Cruz Biotechnology). The membranes were washed and incubated with horseradish peroxidase-coupled goat anti-rabbit IgG (Sigma), peroxidase-coupled rabbit anti-goat IgG and horseradish peroxidase-coupled goat anti-mouse IgG (Sigma) for 45 min and were developed through Western Lightning Chemiluminescence (PerkinElmer Life and Analytical Sciences). Protein expression was calculated as a ratio towards specific total protein expression or β-actin (1:5000; Santa Cruz Biotechnology) detection.
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