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Pakt1 s473

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PAKT1 S473 is a reagent used in cell biology research. It is an antibody that specifically recognizes the phosphorylated form of the AKT1 protein at serine residue 473. This phosphorylation event is important for the activation of the AKT1 protein, which plays a key role in various cellular signaling pathways.

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Lab products found in correlation

Pakt2 s474, by Cell Signaling Technology (2 mentions) Pakt s473, by Cell Signaling Technology (2 mentions) Rad001, by Selleck Chemicals (1 mentions) Pan akt, by Cell Signaling Technology (1 mentions) Ps6 s240 244, by Cell Signaling Technology (1 mentions) Pakt t308, by Cell Signaling Technology (1 mentions) Antibody against hsc 70, by Santa Cruz Biotechnology (1 mentions) Pgsk3α β s21 9, by Cell Signaling Technology (1 mentions) Pher2 y877, by Cell Signaling Technology (1 mentions) Recombinant human tgf β1, by Thermo Fisher Scientific (1 mentions) Recombinant human egf, by Thermo Fisher Scientific (1 mentions) Mmp 2, by Cell Signaling Technology (1 mentions) Sds page, by Thermo Fisher Scientific (1 mentions) Enhanced chemi luminescence (ecl), by Cytiva (1 mentions) Actin, by Merck Group (1 mentions) Human insulin solution, by Merck Group (1 mentions) Horse serum, by Thermo Fisher Scientific (1 mentions) Pierce a g magnetic beads, by Thermo Fisher Scientific (1 mentions) Bovine serum albumin (bsa), by Thermo Fisher Scientific (1 mentions) Human skeletal myoblasts, by Thermo Fisher Scientific (1 mentions)

4 protocols using pakt1 s473

1

Comprehensive Signaling Pathway Analysis

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Antibodies against AKT1, AKT2, AKT3, panAKT, pAKT S473, pAKT T308, pAKT1 S473, pAKT2 S474, pS6 S240/244, pGSK3α/β S21/9, pHER2 Y877, S100A4, MMP2, RANK, and CTGF, and secondary HRP-linked antibodies against rabbit or mouse IgG were purchased from Cell Signaling Technology Inc. (Danvers, MA, USA). Antibody against AKT3 was provided by Millipore (Burlington, MA, USA). Antibody against HSC-70 was purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA). Antibody against pDDR1/2 Y796/Y740 was provided by R&D systems Inc. (Minneapolis, MN, USA). RAD001 was provided by Selleck Chemicals (Houston, TX, USA) and MK-2206 was obtained from AbMole BioScience Inc. (Houston, TX, USA). Recombinant human TGF-β1 and recombinant human EGF were purchased from PeproTech Inc. (Rocky Hill, NJ, USA).
Hinz N., Baranowsky A., Horn M., Kriegs M., Sibbertsen F., Smit D.J., Clezardin P., Lange T., Schinke T, & Jücker M. (2021). Knockdown of AKT3 Activates HER2 and DDR Kinases in Bone-Seeking Breast Cancer Cells, Promotes Metastasis In Vivo and Attenuates the TGFβ/CTGF Axis. Cells, 10(2), 430.
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2

Western Blot Analysis of Signaling Proteins

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For Western blot analysis, 50–100 μg of lysate was separated by SDS-PAGE (Invitrogen) and transferred to nitrocellulose membrane. After blocking in 5% BSA in PBST for one hour, membranes were incubated with the appropriate primary antibody followed by secondary antibody coupled with horseradish peroxidase. The primary antibodies used were against mTOR, pmTOR (S2248), RSK2, pRSK2 (S380), ERK1/2, pERK1 (T202/Y204), AKT1 and pAKT1 (S473), purchased from Cell Signaling Technology (Andover, MA). Actin was purchased from Sigma Aldrich, MO. Custom mouse monoclonal antibodies were made against EGFRL858R, that recognizes mutant EGFR, but not WT EGFR, and against EGFRL858, that recognizes wild type EGFR, but not mutant EGFR in collaboration with NanoTools (Germany). Membranes were incubated with ECL (Amersham) for 5 minutes prior to exposing to X-ray film.
Zhang X., Belkina N., Jacob H.K., Maity T., Biswas R., Venugopalan A., Shaw P.G., Kim M.S., Chaerkady R., Pandey A, & Guha U. (2015). Identifying novel targets of oncogenic EGF receptor signaling in lung cancer through global phosphoproteomics. Proteomics, 15(2-3), 340-355.
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3

Investigating AKT Signaling in Human Skeletal Myoblasts

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Human Skeletal Myoblasts (#A11440 or #A12555), low glucose Dulbecco's Modified Eagle Medium (#11885092), Horse Serum (#26050070), Bovine Serum Albumin (#A9576), and Pierce A/G Magnetic Beads (#88802) were purchased from Thermofisher Scientific Antibodies for Total AKT1 (#2938), Total AKT2 (#2964), Total AKT3 (#14293), Total AKT (#4685), p‐AKT S473 (#4060), p‐AKT1 S473 (#9018), p‐AKT2 S474 (#8599), AKT (Pan) (C67E7) (rabbit monoclonal; #4691), AKT (Pan) (40D4) (mouse monoclonal; #2920), and Human Insulin‐like Growth Factor 1 (#8917) were purchased from Cell Signaling Technologies. Secondary antibodies Anti‐Rabbit IgG HRP‐Linked Antibody (PN#7074) and Anti‐Mouse IgG HRP‐Linked Antibody (PN#7076) were purchased from Cell Signaling Technologies. Human Insulin Solution (#I9278) was purchased from Sigma Aldrich ImageJ was downloaded from the National Institutes of Health website.
Roberts B.M., Geddis A.V, & Matheny RW J.r. (2023). Differential activation of AKT isoforms by growth factors in human myotubes. Physiological Reports, 11(20), e15805.
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4

Western Blot Analysis of Metabolic Proteins

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Tissues designated for western blot were harvested and snap frozen in liquid nitrogen. Tissues were lysed and homogenized as described [26 ]. Modifications to the protocol include a substitution to a 2X RIPA lysis buffer and the addition of two extra centrifugation steps at 10,000g for 5 minutes to reduce the congealed lipid levels found in the liver lysates. Protein concentrations were then quantified with Coomassie Bradford Assay. 20–50 μg of samples were electrophoresed at 4° C and transferred overnight (16 hours, 20V at 4°C). Primary antibodies used are pAMPKT172 (1:2000; Cell Signaling Technology), AMPK (1:2000; Cell Signaling Technology), Glut4 (1:750; Santa Cruz Biotechnology), Glut2 (1:2000, Millipore), HKII (1: 10,000; Millipore), phospho-Glycogen SynthaseS641/S645 (pGS) (1:10,000; Invitrogen), pACC1/2 (1:8,000; Cell Signaling Technology), ACC1/2 (1:8,000; Cell Signaling Technology), OxPhos cocktail (1:2000; AbCam), VDAC (1:7,000; AbCam), pAKT1S473 (1:2000;Cell Signaling Technology), AKT1 (1:2000;Cell Signaling Technology). Bands at predicted sizes were acquired on a ChemiDoc ™ XRS Systems. Optical densities were quantified using Image Lab software and normalized against total protein content acquired through Ponceau S staining or β-actin (1:3000, Sigma Life Science).
Heydemann A., González-Vega M., Berhanu T.K., Mull A.J., Sharma R, & Holley-Cuthrell J. (2016). Hepatic Adaptations to a High Fat Diet in the MRL Mouse Strain are Associated with an Inefficient Oxidative Phosphorylation System. Jacobs journal of diabetes and endocrinology, 2(1), 013.
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