Sirtuin 1

Manufactured by Cell Signaling Technology

Sourced in United States

Sirtuin 1 is a protein that plays a key role in cellular metabolism and longevity. It is an enzyme that regulates the activity of other proteins by removing acetyl groups from them, a process known as deacetylation. Sirtuin 1 is involved in a variety of cellular processes, including DNA repair, gene expression, and the regulation of mitochondrial function.

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Lab products found in correlation

β actin, by Santa Cruz Biotechnology (2 mentions) P pak4, by Cell Signaling Technology (1 mentions) Ecl plus solution, by Thermo Fisher Scientific (1 mentions) Fk866, by Bio-Techne (1 mentions) Goat anti mouse and goat anti rabbit hrp conjugated igg, by Bio-Rad (1 mentions) Anti nampt, by Fortis Life Sciences (1 mentions) C myc, by Cell Signaling Technology (1 mentions) Mouse monoclonal anti β actin, by Merck Group (1 mentions) β catenin, by Cell Signaling Technology (1 mentions) P β catenin, by Cell Signaling Technology (1 mentions) β actin, by Cell Signaling Technology (1 mentions) Mtt solution, by Merck Group (1 mentions) Sunitinib, by LC Laboratories (1 mentions) Angiomotin, by Abcam (1 mentions) Vegfr2, by Abcam (1 mentions) Vegfa, by Abcam (1 mentions) Thrombospondin 1, by Abcam (1 mentions) Anti pgc 1α, by Merck Group (1 mentions) Anti grp78 bip, by Abcam (1 mentions) Caspase 3 casp 3, by Cell Signaling Technology (1 mentions)

5 protocols using sirtuin 1

Protein Expression Quantification Protocol

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MTT solution, mouse monoclonal anti-β-actin, NA, NMN and NAD were obtained from Sigma (St. Louis, MO, USA). The antibodies against (PAK4, p-PAK4, β-catenin, p-β-catenin, cycline D1, c-Myc, β-actin (rabbit), PARP, Sirtuin 1) were from cell signaling Technology, Inc. (Beverly, MA, USA). Goat anti-mouse and goat anti-rabbit HRP conjugated IgG were obtained from Bio-Rad (Hercules, CA). Anti-NAMPT was from Bethyl Laboratories (Montgomery, TX, USA), anti-NAPRT1 was from Proteintech (Rosemont, IL 60018, USA). ECL Plus solution was from Thermo-Fisher Scientific (Waltham MA, USA). KPT-9274 and its vehicle were from Karyopharm Therapeutics (Newton, MA, USA). FK866 was from TOCRIS Biosciences. Sunitinib was obtained from LC laboratories (Woburn, MA).

Aboud O.A., Chen C.H., Senapedis W., Baloglu E., Argueta C, & Weiss R.H. (2016). Dual and specific inhibition of NAMPT and PAK4 by KPT-9274 decreases kidney cancer growth. Molecular cancer therapeutics, 15(9), 2119-2129.

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Characterization of ECFC dysfunction in IUGR

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CTRL-ECFCs (n = 4–5) and IUGR-ECFCs (n = 4) were fixed using cold ethanol 70% and stained with sirtuin-1, endothelial nitric oxide synthase (eNOS), 53BP-1 (rabbit, 1:100; cell signaling, Danvers, MA, USA), CD34, angiopoietin, angiomotin, VEGF-A, VEGFR-2, and thrombospondin-1 (rabbit, 1:100; Abcam, Cambridge, UK) overnight at 4 °C. ECFCs were then washed with PBS and incubated for 2 h with Alexa Fluor-488 goat anti-rabbit IgG (IgG 1:200), Abcam), and were rinsed with PBS and mounted using Fluoromount-G mounting medium with DAPI. Autofluorescence was subtracted. A negative control was obtained using incubation only with the secondary antibody. The slides were observed blindly using a fluorescence microscope (Eclipse Ti2 Series) by the same experimenter. Three images per culture well (2 wells per ECFCs) were captured. Fluorescence of each factor was normalized to DAPI fluorescence. The pictures were evaluated using ImageJ software. Each experiment was performed in duplicate.

Simoncini S., Coppola H., Rocca A., Bachmann I., Guillot E., Zippo L., Dignat-George F., Sabatier F., Bedel R., Wilson A., Rosenblatt-Velin N., Armengaud J.B., Menétrey S., Peyter A.C., Simeoni U, & Yzydorczyk C. (2021). Endothelial Colony-Forming Cells Dysfunctions Are Associated with Arterial Hypertension in a Rat Model of Intrauterine Growth Restriction. International Journal of Molecular Sciences, 22(18), 10159.

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Quantitative Western Blot for Liver Proteins

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Antibodies against rat phosphorylated AMP-activated protein kinase (phospho-AMPK) α and β, total AMPKα and AMPKβ, phosphorylated protein kinase A catalyst unit (phospho-PKA C), sirtuin-1, caspase 3 (CASP3), and GAPDH were purchased from Cell Signaling (Danvers, Mass). Anti-GRP78/BIP and anti–peroxisome proliferator-activated receptor α (PPAR-α) antibodies were purchased from Abcam (Cambridge, Mass). Anti-NLRP3 and anti–PGC-1α antibodies were purchased from EMD Millipore (Billerica, Mass). SuperSignal West Pico chemiluminescent substrate was purchased from Thermo Scientific Inc. (Rockford, Ill).
Approximately 40 mg of frozen liver tissue was homogenized in 150 mmol/L NaCl, 20 mmol/L Tris-HCl (pH 7.5), 1% (wt/vol) NP-40, 1 mmol/L EDTA, 1 mmol/L EGTA, 1 mmol/L sodium orthovanadate, 1 mmol/L β-glycerol phosphate, 2.5 mmol/L sodium pyrophosphate, and 1× complete protease inhibitor mixture (Roche Molecular Biochemicals, Indianapolis, Ind). The homogenate was centrifuged at 12,000g for 30 min at 4°C, and the pellet discarded. Western blotting was performed with 30 µg of protein per well. Band intensities were quantified with the Image J software (National Institutes of Health, Bethesda, Md). GAPDH was used as loading control.

Diao L., Marshall A.H., Dai X., Bogdanovic E., Abdullahi A., Amini-Nik S, & Jeschke M.G. (2014). BURN PLUS LIPOPOLYSACCHARIDE AUGMENTS ENDOPLASMIC RETICULUM STRESS AND NLRP3 INFLAMMASOME ACTIVATION AND REDUCES PGC-1α IN LIVER. Shock (Augusta, Ga.), 41(2), 138-144.

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Immunoblotting Analysis of Aortic and HUVEC Proteins

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Immunoblotting was performed as described previously [40 (link)]. Briefly, thoracic aorta tissue and HUVEC lysates were sonicated, separated on a polyacrylamide gel and transferred onto PVDF membranes. Blocked membranes were proved with primary antibodies against p-eNOS (#9571, Cell Signaling Technologies, USA), eNOS (#32027, Cell Signaling Technologies), nitrotyrosine (#sc32757, Santa Cruz Biotechnologies, USA), p-IRE1α (#ab124945, Abcam, UK), IRE1α (#3294, Cell Signaling Technologies), sXBP-1 (#ab220783, Abcam), GRP78 (#sc166490, Santa Cruz Biotechnologies), CHOP (#2895, Cell Signaling Technologies), Sirtuin 1 (#8469, Cell Signaling Technologies), p-AMPK (#2535, Cell Signaling Technologies), AMPK (#5832, Cell Signaling Technologies), and β-actin (#sc47778, Santa Cruz Biotechnologies). Blots were washed with TBS-T buffer, blocked with 5% Skim milk or BSA during 1 h and probed again with species-specific horseradish peroxidase conjugated secondary antibodies, anti-rabbit HRP 2nd antibody specifically for p-eNOS, eNOS, p-IRE1α, IRE1α, sXBP-1, CHOP, Sirtuin 1, p-AMPK, and AMPK and anti-mouse HRP 2nd antibody specifically for nitrotyrosine, GRP78 and β-actin. Protein signals were developed using ECL (Bio-Rad, Hercules, CA, USA), and their quantifications were performed by measuring the band intensity using the Image J analysis system (NIH, Bethesda, MD, USA).

Lee G.H., Lee H.Y., Lim Y.J., Kim J.H., Jung S.J., Jung E.S., Chae S.W., Lee J., Lim J., Rashid M.M., Min K.H, & Chae H.J. (2023). Angelica gigas extract inhibits acetylation of eNOS via IRE1α sulfonation/RIDD-SIRT1-mediated posttranslational modification in vascular dysfunction. Aging (Albany NY), 15(23), 13608-13627.

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Quantification of Renal Protein Levels

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The protein level in the renal tubule cells and kidney tissues was determined by Western blotting as previously described^{15 (link)}. The sodium dodecyl sulfate (SDS)-PAGE was used to separate equal amount of proteins, and then transferred electrophoretically onto polyvinylidene difluoride (PVDF) membranes. The membranes were incubated with primary antibodies for GRP78 (#3183), Chop (#2895), Bax (#2772), Bcl-2 (#15071), cleaved caspase-3 (#9664), Sirtuin-1 (Sirt1; #8469), phosphorylated NFκB-p65 (#3033) (Cell Signalling, Danvers, MA, USA), catalase (#ab16731), superoxide dismutase 1 (SOD1; #ab13498), Klotho (#ab203576), CD68 (#ab125212) (abcam, Cambridge, MA, USA), NFκB-p65 (#sc-8008), cyclooxygenase (Cox)-2 (#sc1745), β-actin (#sc-47778) (Santa Cruz, Dallas, TX, USA), inducible nitric oxide synthase (iNOS; #610329) (BD, Franklin Lakes, NJ, USA), and Ly6G (#14-5931-82) (eBioscience, San Diego, CA), and then incubated with horseradish peroxidase-conjugated secondary antibodies (Bio-Rad, Hercules, CA, USA). The signals were detected by using enhanced chemiluminescence substrates (Bio-Rad), and developed with a Fuji Blue X-Ray Film. Protein bands were quantitated by using ImageJ software. The raw data/full blot summary is available in the Supplementary Fig. 5.

Chiang C.K., Loh J.Z., Yang T.H., Huang K.T., Wu C.T., Guan S.S., Liu S.H, & Hung K.Y. (2020). Prevention of acute kidney injury by low intensity pulsed ultrasound via anti-inflammation and anti-apoptosis. Scientific Reports, 10, 14317.

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