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Phospho ampkα t172

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Phospho-AMPKα T172 is a primary antibody that specifically recognizes the phosphorylated form of the AMPK alpha subunit (AMPKα) at threonine 172. AMPK is a key cellular energy sensor that plays a crucial role in regulating metabolic pathways.

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Lab products found in correlation

Ampkα, by Cell Signaling Technology (6 mentions) 4e bp1, by Cell Signaling Technology (3 mentions) Phospho ulk1 s757, by Cell Signaling Technology (2 mentions) Phospho stat3 tyr705, by Cell Signaling Technology (1 mentions) Phospho stat3 ser727, by Cell Signaling Technology (1 mentions) Alexa fluor 488 donkey anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Vimentin 10366 1 ap, by Proteintech (1 mentions) E cadherin 20874 1 ap, by Proteintech (1 mentions) Phospho 4ebp1, by Cell Signaling Technology (1 mentions) P70s6k, by Cell Signaling Technology (1 mentions) Phospho p70s6k, by Cell Signaling Technology (1 mentions) Caveolin 1, by Cell Signaling Technology (1 mentions) Phospho enos ser1177, by Cell Signaling Technology (1 mentions) Anti hur, by Merck Group (1 mentions) Histone h3, by Cell Signaling Technology (1 mentions) Anti cd31, by Abcam (1 mentions) Phospho gsk3β s9, by Cell Signaling Technology (1 mentions) Lxrα β, by Santa Cruz Biotechnology (1 mentions) Phospho acc1, by Cell Signaling Technology (1 mentions) Ampkβ1, by Cell Signaling Technology (1 mentions)

Spelling variants of this product

AMPKα (280 citations) Phospho-AMPKα (99 citations) Rabbit anti-phospho-AMPKα T172 (3 citations) Anti-phospho-T172 AMPKα ( (2 citations)

9 protocols using phospho ampkα t172

1

Comprehensive Antibody Panel for Cellular Analysis

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Antibodies to RKIP (CST#5291), AMPKα (CST#5832), phospho-AMPKα (T172) (CST#8208), p70S6K (CST#2708), phospho-p70S6K (CST#9208), 4EBP1 (CST#9644), phospho-4EBP1 (CST#9459), STAT3 (CST#9132), phospho-STAT3 (Tyr705) (CST#9131) and phospho-STAT3 (Ser727) (CST#9134) were purchased from Cell Signaling Technology. Antibodies to E-Cadherin (20874-1-AP) and vimentin (10366-1-AP) were obtained from Proteintech Group. Anti-ANXA7 antibody (sc-17815) used for co-immunoprecipitation and anti-p-Thr antibody (sc-5267) were obtained from Santa Cruz Biotechnology. Anti-ANXA7 antibody (A4475 MSDS) used for western blot and antibody to β-actin (A1978 MSDS) were purchased from Sigma. Horseradish peroxidase-conjugated secondary antibodies for Western blot were from Santa Cruze Biotechnology. Secondary antibody for immunofluorescence was donkey anti-rabbit IgG Alexa Fluor-488 (CA21206s, Invitrogen).
Liu S., Li X., Lin Z., Su L., Yan S., Zhao B, & Miao J. (2017). SEC-induced activation of ANXA7 GTPase suppresses prostate cancer metastasis. Cancer letters, 416, 11-23.
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2

Antibody Validation for AMPK Signaling

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Antibodies to caveolin-1, phospho-AMPKα T172, AMPKα, Histone H3, eNOS, and phospho-eNOS Ser1177 were from Cell Signaling Technology (Danvers, MA). Antibodies to LKB1 and GAPDH were purchased from Santa Cruz Biotechnology (Santa. Cruz, CA). Anti-HuR was purchased from Millipore (Temecula, CA). Anti-CD31 was from Abcam (Cambridge, UK). FuGene HD transfection reagent was from Roche Applied Science (Indianapolis, IN).
Zhang W., Wang Q., Wu Y., Moriasi C., Liu Z., Dai X., Wang Q., Liu W., Yuan Z.Y, & Zou M.H. (2014). Endothelial Cell-Specific LKB1 Deletion Causes Endothelial Dysfunction and Hypertension in Mice in vivo. Circulation, 129(13), 1428-1439.
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3

Immunoprecipitation and Immunoblotting Protocol

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For immunoprecipitation, cells were lysed in IP buffer (150mM NaCl, 50mM Tris pH7.5, 1% Triton X-100) clarified by centrifugation at 14,000 rpm for 15 minutes, then incubated with primary antibody for 2 hours and then incubated with Protein A-Agarose beads (Roche) for 30 min before washing in IP buffer 3 times. Protein samples for immunoblotting were lysed in 1x Lamelli buffer and subjected to SDS-PAGE with a tris-glycine buffer system. Rabbit antibodies against phospho-GSK3β(S9), phospho-S6K(T389), S6K, phospho-AKT(T308), phospho-AKT(T473), AKT, phospho-AMPKα(T172), AMPKα, AMPK β1, phospho-ACC1, ACC1, phospho-TBC1D1(S700), TBC1D1, HSP90, YAP, TSC1, and Rpl26 were from Cell Signaling. PPARα antibody (3B6/PPAR) was purchased from Alexis (Now part of Enzo life sciences). LXRα/β was from Santa Cruz. Mouse antibody anti-SREBP-1 was from BD Biosciences. Anti-HA antibody was from Roche. Anti-FLAG(M2) was from Sigma. Monoclonal mouse anti-Actin was from Developmental Studies Hybridoma Bank. Anti-PPP2R5C was generated by immunizing guinea pigs with NCBI Variant 3 of mouse PPP2R5C.
Cheng Y.S., Seibert O., Klöting N., Dietrich A., Straßburger K., Fernández-Veledo S., Vendrell J.J., Zorzano A., Blüher M., Herzig S., Berriel Diaz M, & Teleman A.A. (2015). PPP2R5C Couples Hepatic Glucose and Lipid Homeostasis. PLoS Genetics, 11(10), e1005561.
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4

Drosophila Proteasome and Signaling Antibodies

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The polyclonal antibody against the Drosophila β5 proteasome subunit was a gift from Prof. M. Figueiredo-Pereira (Department of Biological Sciences, Hunter College of the City University of New York, NY). Primary antibodies against the 20S-α (sc-65755), Rpn10 (sc-65748), and Rpn7 (sc-65750) Drosophila proteasome subunits; the anti-Hsp70 (sc-25837), anti-ubiquitin (Ub) (sc-8017), anti-AMPKα (sc-25792), anti-GAPDH (sc-25778); anti-β-tubulin (sc-20852) and the HRP-conjugated secondary antibodies were from Santa Cruz Biotechnology (Heidelberg, Germany). The antibodies against Gsk-3 (05-412) and the Rpn6 (NBP1-46191) proteasome subunit were obtained from Millipore S.A. and Novus Inc. respectively. The antibodies against phospho-Gsk-3S21/S9 (#9331), phospho-AktS505 (#4054), Akt (#9272), phospho-AMPKαT172 (#2535), and the phosphoSer/Thr Pdpk1 docking motif (#9634) were purchased from Cell Signaling; the anti-AGE antibody was from Cosmo Bio (KAL-KH001-01).
Tsakiri E.N., Gaboriaud-Kolar N., Iliaki K.K., Tchoumtchoua J., Papanagnou E.D., Chatzigeorgiou S., Tallas K.D., Mikros E., Halabalaki M., Skaltsounis A.L, & Trougakos I.P. (2017). The Indirubin Derivative 6-Bromoindirubin-3′-Oxime Activates Proteostatic Modules, Reprograms Cellular Bioenergetic Pathways, and Exerts Antiaging Effects. Antioxidants & Redox Signaling, 27(14), 1027-1047.
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5

Antibody Validation for Autophagy Signaling

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Phospho-AMPKα T172 (2531; 1:1000), AMPKα1 (2532; 1:1000), phospho-ULK1 S757 (14202; 1:1000), phospho-beclin 1 S15 (13825; 1:1000), beclin 1 (3495; 1:1000), PKCα (2056; 1:1000), ULK1 (8054; 1:1000), ubiquitin (3933; 1:1000), 4E-BP1 (9452; 1:1000), phospho-4E-BP1 Thr37/46 (2855; 1:1000), mTOR (2983; 1:1000), phospho-mTOR S2448 (5536; 1:1000), phospho-(Ser/Thr) PKC substrate (6967; 1:1000), β-tubulin (2146; 1:3000), HA (2999; 1:3000), GFP (2956; 1:2000), and Myc (2278; 1:2000) antibodies were from Cell Signaling Technology, Danvers, MA. LC3 antibody (NB100-2220; 1:7000) was from NOVUS Biologicals, Littleton, CO. p-Atg13 S318 (600-401-C49S; 1:1000) was from Rockland Immunochemicals, Limerick, PA. β-Actin antibodies (sc-47778; 1:10000) were from Santa Cruz Biotechnology, Dallas, Texas. P62 (SQSTM1) antibody (ab56416; 1:4000), SNAP29 (ab56566; 1:1000), LAMP2a (ab18528; 1:3000), and VAMP8 (EP2629Y; 1:1000) were from Abcam, Cambridge, MA. GST (E022040-01; 1:10000) was from EARTHOX Life Sciences, Millbrae, CA, and Flag (SLBJ7864V; 1:2000) and STX17 (HPA001204; 1:1000) were purchased from SIGMA-Aldrich, St. Louis, MO. FIP200 (17250-1-AP; 1:1000) was from Proteintech, Rosemont, IL. Total p-S, T (612549; 1:4000) antibody was from BD Biosciences, San Jose, CA. Anti-phospho ULK1 S423 antibody (1:100) was generated and affinity purified by ABclonal Technology, Wuhan, China.
Wang C., Wang H., Zhang D., Luo W., Liu R., Xu D., Diao L., Liao L, & Liu Z. (2018). Phosphorylation of ULK1 affects autophagosome fusion and links chaperone-mediated autophagy to macroautophagy. Nature Communications, 9, 3492.
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6

Protein extraction and immunoblotting protocol

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Cells were lysed in 50 mM Tris–HCl (pH 7.5), 1 mM EGTA, 1 mM EDTA, 1 mM sodium orthovanadate, 50 mM sodium fluoride, 1 mM sodium pyrophosphate, 0.27 M sucrose, 1% (v/v) Triton X-100, 0.1% (v/v) 2-mercaptoethanol, and cOmplete EDTA-free Protease Inhibitor Cocktail tablets (Roche). Lysates were clarified by centrifugation (20800 g for 10 min at 4 °C) and supernatants snap-frozen and stored at −80 °C. Protein concentration was determined with Coomassie Protein Assay Reagent (Thermo Scientific). Proteins were separated on 4–12% gradient Bis-Tris polyacrylamide gels (Invitrogen), and immunoblotting carried out using standard techniques. Antibodies recognising total acetyl-CoA carboxylase (Cat #3676), total AMPKα (#5832), phospho-acetyl-CoA carboxylase S79 (#11818), phospho-AMPKα T172 (#2535), and GAPDH (#5174) were from Cell Signaling Technology and used at a dilution of 1 in 1000. Anti-rabbit horseradish peroxidase (#7074) was from Cell Signaling Technology.
Phair I.R., Nisr R.B., Howden A.J., Sovakova M., Alqurashi N., Foretz M., Lamont D., Viollet B, & Rena G. (2022). AMPK integrates metabolite and kinase-based immunometabolic control in macrophages. Molecular Metabolism, 68, 101661.
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7

Fractionation and Western Blot Analysis

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Cardiomyocytes and isolated heart tissues were homogenized in lysis buffer (Cell Signaling Tech.) containing protease and phosphatase inhibitors. Fractionated cytoplasmic and nuclear protein lysates (20–40 μg) were separated and applied to SDS-PAGE and transferred onto PVDF membrane (BioRad). Antibodies used were phospho- or total against AMPKα, phospho-AMPKα-T172, (Cell Signaling Technology), histone 3 (H3) (Genetex), NFATc and β-actin (Santa Cruz Biotechnology) and resistin (Millipore). Serca2a antibody is custom made in our lab). β-actin expression verified cytosolic protein loading while H3 served as nuclear specific internal control.
Singh R., Moreno P., Hajjar R.J, & Lebeche D. (2018). A role for calcium in resistin transcriptional activation in diabetic hearts. Scientific Reports, 8, 15633.
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8

Western Blot Analysis of Cellular Signaling

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Cells were lysed in buffer containing 50 mM Tris pH 7.4, 1% Nonidet P-40, 0.25% sodium deoxycholate, 1 mM EDTA, 150 mM NaCl, 1 mM dithiothreitol, 1 mM sodium orthovanadate, 20 mM sodium fluoride, 10 mM beta-glycerophosphate, 10 mM sodium pyrophosphate, 2 ug ml−1 aprotinin, 2 ug ml−1 leupeptin and 0.7 ug ml−1 pepstatin. Western blot analysis was performed using standard protocols, and the following commercial antibodies were used as probes: ASNS (Proteintech 14681-1-AP, 1:1000), ATF4 (Cell Signaling 11815, 1:500), phospho-T389 S6 kinase (Cell Signaling 9234, 1:500), S6 kinase (Cell Signaling 2708, 1:1000), phospho-S235/235 S6 ribosomal protein (Cell Signaling 4858, 1:3000), S6 ribosomal protein (Cell Signaling 2217, 1:1000), phospho-S1859 CAD (Cell Signaling 70307, 1:500), CAD (Cell Signaling 11933, 1:1000), 4E-BP1 (Cell Signaling 9644, 1:500), phospho-S757 ULK1 (Cell Signaling 14202, 1:1000), ULK1 (Cell Signaling 8054, 1:1000), phospho-T172 AMPKα (Cell Signaling 2535, 1:500), AMPKα (Cell Signaling 2532, 1:1000), PHGDH (Cell Signaling 13428, 1:500), β-Actin (Cell Signaling 4970, 1:1000), PSAT1 (Novus 89-004-606, 1:500), and a-tubulin (Sigma T6074, 1:10,000).
Krall A.S., Mullen P.J., Surjono F., Momcilovic M., Schmid E.W., Halbrook C.J., Thambundit A., Mittelman S.D., Lyssiotis C.A., Shackelford D.B., Knott S.R, & Christofk H.R. (2021). Asparagine couples mitochondrial respiration to ATF4 activity and tumor growth. Cell metabolism, 33(5), 1013-1026.e6.
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9

Antibody Panel for Angiogenic Signaling

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The following antibodies were from Cell Signalling Technology (Frankfurt, Germany): Total VEGFR2 (#2479), phospho(Y1175)-VEGFR2 (#2478), total VEGFR1 (#64,094), phospho(S1177)-eNOS (#9570), total Akt (#4691), phospho(S473)-Akt (#9271), total Erk (#9107), phospho(T202/Y204)-Erk (#9106), total AMP-activated protein kinase α (AMPKα) (#2532), phospho(T172)-AMPKα (#2531), total p38 (#9212), phospho(T180/Y182)-p38 (#4511), phospho(Y207)-Crk-L (#3181), phospho(Y1086)-EGFR (#2220), total EGFR (#2232). Total eNOS antibody (#610,297) and total ABL-1 antibody were purchased from BD Biosciences (Heidelberg, Germany). Total ABL-2 (#sc-81,154) and total Crk-L (#sc-365,092) antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX, US). Total FRK antibody (#A95,485) was from Antibodies.com (Stockholm, Sweden). Horseradish peroxidase (HRP)-conjugated secondary antibodies to rabbit or mouse IgG were from Kirkegaard and Perry Laboratories, Inc. (Gaithersburg, MD, USA).
Zibrova D., Ernst T., Hochhaus A, & Heller R. (2024). The BCR::ABL1 tyrosine kinase inhibitors ponatinib and nilotinib differentially affect endothelial angiogenesis and signalling. Molecular and cellular biochemistry.
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