Recombinant mouse leptin was purchased from R&D Systems (Minneapolis, MN, USA). Salubrinal was purchased from Calbiochem (La Jolla, CA, USA). Antibodies used in these experiments include anti-phospho-JAK2 Tyr1007/1008 (1:1,000, #3771), anti-phospho-Stat3 Tyr705 (1:1,000, #9145), anti-Stat3 (1:1,000, #8768), anti-phospho-PERK Thr980 (1:1,000, #3179), anti-PERK (1:1,000, #5683), anti-CHOP (1:1,000, #2895), anti-NF-κB p65 (1:1,000, #8242), anti-Lamin B1 (1:1,000, #13435), anti-phospho-IκBα (1:1,000, #2859), anti-IκBα (1:1,000,#9242), and anti-β-Actin (1:5,000, #3700) from Cell Signaling Technology (Beverly, MA, USA). PA was purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). PA was solubilized in pre-heated 0.1 N NaOH and diluted in pre-warmed 10% fatty acid free-BSA solution to give a final concentration of 5 mM. Control media were prepared with 0.1 N NaOH and 10% BSA without lipid. PA solution was freshly made before each experiment.
Anti phospho perk thr980
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-phospho-PERK (Thr980) is a lab equipment product that detects phosphorylation of the PERK protein at threonine 980. It is intended for use in research applications.
Automatically generated - may contain errors
Lab products found in correlation
Anti p erk, by Cell Signaling Technology (3 mentions)
Anti β actin, by Cell Signaling Technology (2 mentions)
Anti bip, by Cell Signaling Technology (2 mentions)
Anti eif2α, by Cell Signaling Technology (2 mentions)
Anti akt, by Cell Signaling Technology (2 mentions)
Recombinant mouse leptin, by R&D Systems (1 mentions)
Anti chop, by Cell Signaling Technology (1 mentions)
Anti phospho stat3 tyr705, by Cell Signaling Technology (1 mentions)
Anti phospho iκbα, by Cell Signaling Technology (1 mentions)
Anti phospho jak2 tyr1007 1008, by Cell Signaling Technology (1 mentions)
Anti stat3, by Cell Signaling Technology (1 mentions)
Anti lamin b1, by Cell Signaling Technology (1 mentions)
Anti nf κb p65, by Cell Signaling Technology (1 mentions)
Anti iκbα, by Cell Signaling Technology (1 mentions)
Anti ire1α, by Cell Signaling Technology (1 mentions)
Anti sp1, by Cell Signaling Technology (1 mentions)
Anti phospho eif2α, by Cell Signaling Technology (1 mentions)
Bca protein estimation assay, by Thermo Fisher Scientific (1 mentions)
Anti p70s6k, by Cell Signaling Technology (1 mentions)
Chloroquine c6628, by Merck Group (1 mentions)
5 protocols using anti phospho perk thr980
1
Palmitic Acid Signaling Pathways
2
Protein Expression Profiling in Treated Cells
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Proteins from treated and untreated cell lysates were estimated using the BCA protein estimation assay (Thermo Scientific). Blots were probed for: anti-Sp1; anti-BiP; anti-Ire1α; anti-PERK; anti-phospho-PERK (Thr980); anti-eIF2α; anti-phospho-eIF2α; anti-β-actin (all from Cell Signaling).
3
Cellular Protein Expression Analysis
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The following primary antibodies were obtained from Cell Signaling Technology (Beverly, MA, USA): anti-BiP (#3177), anti-LC3A/B (#4108), anti-Phospho-p70S6K (Thr389) (#9205), anti-p70S6K (#9202), anti-Phospho-ULK1 (Ser757) (#14202), anti-ULK1 (#6439), anti-phospho-PERK (Thr980) (#3179), anti-PERK (#3192), anti-Phospho-eIF2-alpha (Ser51) (#9721), anti-eIF2-alpha (#9722) anti-Tuberin/TSC2 (#4308). Anti-β-actin and anti-α-tubulin were obtained from Sigma-Aldrich (St. Louis, MO, USA). Anti-p62 (GP62-C-WBC) was obtained from ProGen (Heidelberg, Germany). Anti-Lamin A/C/B1 (EPR4068) was obtained from Abcam. Anti-prelamin A (#MABT345) was obtained from Millipore. Anti-p27 Kip1/CDKN1B (sc-1641) was obtained from Santa Cruz Biotechnology. The following secondary antibodies HRP-conjugated were used: anti-Rabbit (NA934) and anti-Mouse (NA931) were obtained from GE Lifesciences (Marlborough, MA, USA). Anti-Guinea pig HRP-conjugated (90001) was purchased by ProGen (Heidelberg, Germany). Chloroquine (C6628) was obtained from Sigma-Aldrich (St. Louis, MO, USA).
4
Quantitative Immunoblot Analysis of Cellular Signaling
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Muscle lysates were prepared in RIPA buffer (50-mM Tris-HCl, 150-mM NaCl, 1% NP-40, 0.1% SDS, and 0.5% sodium deoxycholate) containing protease inhibitor cocktail (Nacalai Tesque) and phosphatase inhibitor cocktail (Biotool) using a Polytron homogenizer (Kinematica). The protein concentrations of lysates were measured using the bicinchoninic acid method, and immunoblot analyses were performed as described previously [12 (link)]. Proteins were detected using anti-PERK, anti-phospho-PERK (Thr980), anti-eIF2α, anti-phospho-eIF2α (Ser51), anti-Akt, anti-phospho-Akt (Ser473), anti-p70 S6 kinase (S6K), anti-phospho-S6K (Thr389), anti-eukaryotic translation initiation factor 4E (eIF4E)-binding protein (4EBP) 1, and anti-phospho-4EBP1 (Thr37/46), which were purchased from Cell Signaling Technology. GAPDH, which was purchased from MBL international, was used as a loading control. Bands were detected using WesternSure ECL Substrate (Li-Cor Biosciences), and images were acquired using an EZ-Capture II Cooled CCD Camera System (ATTO Corp.). The band intensity was determined using ImageJ software.
5
Investigating TM-25659's Molecular Mechanisms
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TM-25659 was obtained from the Korea Research Institute of Chemical Technology (Korea). Bovine serum albumin (BSA), insulin, PA, and methyl cellulose were purchased from Sigma-Aldrich (USA). Anti-AKT, anti-phospho-AKT (ser473), anti-eIF2α, anti-phospho-eIF2α (ser51), anti-PERK, anti-phospho-PERK (Thr980), and anti-ATF4 antibodies were acquired from Cell Signaling Technology (USA). Anti-GCN2, anti-phospho-GCN2 (Thr898) antibodies were acquired from Biorbyt (UK). Anti-FGF21 antibodies were purchased from Abcam (USA). Anti-actin antibody was obtained from Santa Cruz Biotechnology (USA). Culture media, culture supplements, horse serum, and fetal bovine serum (FBS) were purchased from Gibco-BRL (USA). TM-25659 was dissolved in 0.5% methyl cellulose in water for in vivo administration.
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