Flies were lysed in radioimmunoprecipitation assay buffer (Thermo Fisher Scientific) and proteins were then separated by SDS‐PAGE using standard procedures (Wang et al., 2005 (link)). The antibodies used were mouse anti‐AMPK (Abcam), rabbit anti‐phospho‐AMPK (Cell Signaling), rabbit anti‐phospho‐Drosophila p70 S6 Kinase (Cell Signaling), and mouse anti‐α Tubulin (GeneTex). Protein signals were detected with horseradish peroxidase‐conjugated secondary antibodies and ECL reagent (Thermo Fisher Scientific). Immunoblots were quantified using Image J software.
Rabbit anti phospho ampk
Manufactured by Cell Signaling Technology
Rabbit anti‐phospho‐AMPK is a primary antibody that recognizes the phosphorylated form of AMP-activated protein kinase (AMPK). AMPK is a cellular energy sensor that plays a crucial role in regulating cellular metabolism in response to changes in the AMP/ATP ratio. This antibody can be used to detect and quantify the phosphorylated, active form of AMPK in various experimental systems.
Automatically generated - may contain errors
Lab products found in correlation
Ecl reagent, by Thermo Fisher Scientific (2 mentions)
Radioimmunoprecipitation assay buffer, by Thermo Fisher Scientific (2 mentions)
Mouse anti α tubulin, by GeneTex (2 mentions)
Mouse anti ampk, by Abcam (1 mentions)
Alexa fluor 488 affinipure donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions)
Alexa fluor 488 affinipure donkey anti chicken igy, by Jackson ImmunoResearch (1 mentions)
Mouse anti α tubulin, by Merck Group (1 mentions)
Rabbit anti gst, by Cell Signaling Technology (1 mentions)
Mouse anti rfp, by Proteintech (1 mentions)
Rabbit anti actin, by Merck Group (1 mentions)
Rabbit anti lc3b, by Cell Signaling Technology (1 mentions)
Mitoprofile total oxphos human wb antibody cocktail, by Abcam (1 mentions)
Mouse anti iκbα, by Cell Signaling Technology (1 mentions)
Rabbit anti ampk, by Cell Signaling Technology (1 mentions)
Rabbit anti p62, by Cell Signaling Technology (1 mentions)
Polyvinylidene difluoride membrane, by Roche (1 mentions)
Rabbit anti p65, by Cell Signaling Technology (1 mentions)
Goat anti β actin, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti beclin 1, by Cell Signaling Technology (1 mentions)
Goat anti gapdh, by Santa Cruz Biotechnology (1 mentions)
5 protocols using rabbit anti phospho ampk
1
AMPK and S6 Kinase Immunoblotting
2
Immunostaining and Immunoblotting Antibodies
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Primary antibodies used in this study for immunofluorescence are the following: rabbit anti-vasa (1:5000; R. Lehmann), guinea pig anti–phospho–myosin II (1:1000; R. Ward), chicken anti-vasa (1:500; R. Lehmann), and rabbit anti-RhoGEF2 (1:2500; J. Grosshans).
Secondary antibodies used in this study for immunofluorescence are the following: Cy3 AffiniPure Donkey Anti-Rabbit IgG (immunoglobulin G) (Jackson ImmunoResearch, 711-165-152), Cy3 AffiniPure Donkey Anti-Guinea Pig IgG (Jackson ImmunoResearch, 706-165-148), Alexa Fluor 488 AffiniPure Donkey Anti-Rabbit IgG (Jackson ImmunoResearch, 711-545-152), and Alexa Fluor 488 AffiniPure Donkey Anti-Chicken IgY (Jackson ImmunoResearch, 703-545-155).
Primary antibodies used for immunoblotting are the following: mouse anti-mNeongreen (1:1000; Chromotek, 32F6), rabbit anti-mNeongreen (1:1000; Cell Signaling Technology, 53061), mouse anti–α-tubulin (1:4000; Sigma-Aldrich, T6199), mouse anti-RFP (1:1000; Chromotek, 6G6), rabbit anti-GST (1:2000; Cell Signaling Technology, 2622), rabbit anti–phospho-AMPK (1:1000; Cell Signaling Technology, 2535), and mouse anti-AMPK (1:3000; Bio-Rad, MCA2672GA).
Secondary antibodies used in this study for immunoblotting are the following: horseradish peroxidase (HRP) goat anti-rabbit IgG (1:10,000; Abcam, ab6721) and HRP rabbit anti-mouse IgG (1:10,000; Abcam, ab6728).
Secondary antibodies used in this study for immunofluorescence are the following: Cy3 AffiniPure Donkey Anti-Rabbit IgG (immunoglobulin G) (Jackson ImmunoResearch, 711-165-152), Cy3 AffiniPure Donkey Anti-Guinea Pig IgG (Jackson ImmunoResearch, 706-165-148), Alexa Fluor 488 AffiniPure Donkey Anti-Rabbit IgG (Jackson ImmunoResearch, 711-545-152), and Alexa Fluor 488 AffiniPure Donkey Anti-Chicken IgY (Jackson ImmunoResearch, 703-545-155).
Primary antibodies used for immunoblotting are the following: mouse anti-mNeongreen (1:1000; Chromotek, 32F6), rabbit anti-mNeongreen (1:1000; Cell Signaling Technology, 53061), mouse anti–α-tubulin (1:4000; Sigma-Aldrich, T6199), mouse anti-RFP (1:1000; Chromotek, 6G6), rabbit anti-GST (1:2000; Cell Signaling Technology, 2622), rabbit anti–phospho-AMPK (1:1000; Cell Signaling Technology, 2535), and mouse anti-AMPK (1:3000; Bio-Rad, MCA2672GA).
Secondary antibodies used in this study for immunoblotting are the following: horseradish peroxidase (HRP) goat anti-rabbit IgG (1:10,000; Abcam, ab6721) and HRP rabbit anti-mouse IgG (1:10,000; Abcam, ab6728).
3
Mitochondrial Protein Analysis by SDS-PAGE and Western Blot
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SDS-PAGE and Western blots were performed using 20 ug of total protein lysate as previously described [46 , 49 (link)]. Antibodies used were: rabbit anti-LC3B (Cell Signaling Technology, Danvers, MA), rabbit anti-phospho AMPK (Cell Signaling Technology, Danvers, MA), rabbit anti-actin (Sigma Aldrich, St. Louis, MI), Mitoprofile Total OXPHOS Human WB antibody cocktail (Abcam, Cambridge, MA), rabbit anti-pyruvate dehyrogenase (Cell Signaling Technology), rabbit anti phospho-pyruvate dehyrogenase E1 (S293) (Abcam) and anti-rabbit UCP2 (Abcam).
4
Western Blot Analysis of Cardiac Markers
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Western blot analysis was performed as we described previously [18] (link). The border zone of the infarct hearts was separated and homogenized. Equal amounts of protein (30 mg) were separated on SDS–polyacrylamide gels (10%) and electrotransferred to polyvinylidene difluoride membranes (Roche, Switzerland). The membranes were incubated with rabbit anti-LC3 (1∶1000 dilution; Cat#L7543, Sigma), rabbit anti-Beclin-1 (1∶1000 dilution; Cat#3738, Cell signaling technology), rabbit anti-p65 (1∶1000 dilution; Cat#9936S, Cell signaling technology), mouse anti-IκBα (1∶1000 dilution; Cat#9936S, Cell signaling technology), rabbit anti-P62(Cat#5114 Cell signaling technology), rabbit anti-AMPK (Cat#5831, Cell signaling technology), rabbit anti-Phospho-AMPK (Cat#2535, Cell signaling technology), mouse anti-HIF-1α (NB100-105, Novus Biologicals), goat anti-GAPDH and goat anti-β-actin (1∶1000 dilution; Cat#SC-48166 and Cat#SC-1616, Santa Cruz Biotechnology) at 4°C overnight, and incubated with either goat anti-rabbit, rabbit anti-mouse or mouse anti-goat second antibody (1∶5000 dilution; Santa Cruz Biotechnology) for 1 hour at room temperature. Blots were developed using a chemiluminescent substrate and molecular band intensity was determined by densitometry.
5
Immunoblotting of Phospho-AMPK Signaling
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Cells were lysed in radioimmunoprecipitation assay buffer (Thermo), and proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride membranes (Invitrogen), using standard procedures (Huang et al., 2015) . The antibodies used were rabbit anti-phospho-AMPK (1:1000, Cell Signaling), rabbit anti-AMPK (1:500, Abcam) and mouse anti-αtubulin (1:500, GeneTex). Protein signals were visualized with horseradish peroxidase-conjugated secondary antibodies and ECL reagent (Thermo). The intensity of each target protein band was quantified using Image J software.
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