Acetylated tubulin t7451

Manufactured by Merck Group

Sourced in United States

Acetylated tubulin (T7451) is a laboratory reagent used for the detection and analysis of acetylated microtubules in cells. It is a modified form of the tubulin protein, which is a major component of the cytoskeleton in eukaryotic cells. The acetylation of tubulin can influence microtubule stability and function, and is often used as a marker for certain cellular processes.

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Acetylated tubulin (77 citations) T7451 (65 citations) Anti-acetylated α-tubulin (T7451) (3 citations)

6 protocols using acetylated tubulin t7451

Antibody Immunofluorescence and Western Blot

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Mouse monoclonal antibodies: acetylated Tubulin (T7451, IF: 1:10,000; Sigma-Aldrich), ARL13B (66739-1-Ig, IF: 1:300; Proteintech, 75-287, IF: 1:1,000; NeuroMab), and S-tag (MAC112, IF: 1:100, WB: 1:5,000; EMD Millipore). Rat monoclonal antibody: HA (7c9, WB: 1:1,000; ChromoTek). Rabbit polyclonal antibodies: EGFP (50403-2-AP, IF: 1:200, WB: 1:1,000; Proteintech), Flag (F7425, WB: 1:1,000; Sigma-Aldrich), Adenylyl cyclase III (C-20, IF: 1:100; Santa Cruz), B9D1 (NBP2-84489, WB: 1:1,000; Novus Biologicals) and HTR6 (31 (link)). Goat polyclonal antibodies: γ-Tubulin (sc-7396, IF: 1:200; Santa Cruz). Alexa Fluor (AF)–conjugated donkey secondary antibodies from Thermo Fisher Scientific (all used for IF at 1:10,000): AF488 anti-mouse IgG (A21202), AF488 anti-rabbit IgG (A21206), AF555 anti-mouse IgG (A31570), AF555 anti-rabbit IgG (A31572), AF594 anti-rabbit IgG (A21207), and AF647 anti-goat IgG (A21447). Also from Thermo Fisher Scientific were HRP-conjugated secondary antibodies (used for WB at 62 ng/ml): goat anti-rabbit IgG (A16104), goat anti-mouse IgG (A16072) and donkey anti-rat IgG (A18739). Biotinylated proteins were detected with Neutravidin-HRP (A2664, 1 μg/ml; Thermo Fisher Scientific). D-biotin was from Thermo Fisher Scientific (BP-232-1).

Barbeito P., Tachibana Y., Martin-Morales R., Moreno P., Mykytyn K., Kobayashi T, & Garcia-Gonzalo F.R. (2020). HTR6 and SSTR3 ciliary targeting relies on both IC3 loops and C-terminal tails. Life Science Alliance, 4(3), e202000746.

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Poly(I:C) Transfection and Immunostaining

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Polyinosinic:polycytidylic acid [Poly(I:C)], MTT [3(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide] and mouse antibodies which are reactive with acetylated tubulin (T7451) were from Sigma-Aldrich Co (St. Louis, MO, USA). Polyethylenimine (PEI) was purchased from Polysciences, Inc (Warrington, PA, USA). Rabbit antibodies which are reactive with Flag (14793) were from Cell Signaling Technology Inc (Danvers, MA, USA). Rabbit antibodies which are reactive with Arl13b (17711-1-AP) were from Proteintech Group Inc (Rosemont, IL, USA). Goat anti-rabbit-Alexa 568 (A-11011) was obtained from Invitrogen (Calsbad, CA, USA). Human pCMV3-ACE2-Flag plasmid (HG10108-CF) and SARS-CoV-2 (2019-nCoV) spike protein (40592-V08B-B) were purchased from Sino Biological Inc (Wayne, PA, USA). Fibronectin, bovine collagen type I and bovine serum albumin (BSA) were purchased from Advanced BioMatrix (Carlsbad, CA, USA) Except where indicated, all other materials are obtained from the Sigma Chemical Co.

Gweon B., Jang T.K., Thuy P.X, & Moon E.Y. (2022). Primary Cilium by Polyinosinic:Polycytidylic Acid Regulates the Regenerative Migration of Beas-2B Bronchial Epithelial Cells. Biomolecules & Therapeutics, 30(2), 170-178.

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Nrf2, Tubulin, and Cilia Imaging Assay

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N-acetyl-L-cysteine(NAC), hydrogen peroxide(H₂O₂), MTT [3(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide] and 4′,6-diamidno-2-phenylinole(DAPI) were purchased from the Sigma Chemical Co. (St. Louis, MO, USA). 2’,7’–dichlorofluorescin diacetate (DCF-DA) was purchased from Molecular Probe (Eugene, Oregon, USA). Rabbit antibodies which are reactive with Nrf2 (12,721) were from Cell Signaling Technology Inc. (Danvers, MA, USA). Mouse antibodies which are reactive with acetylated tubulin (T7451) or β-tubulin (T4026) were from Sigma-Aldrich Co. (St. Louis, MO, USA). Rabbit antibodies which are reactive with Arl13b (17,711–1-AP) were from Proteintech Group Inc. (Rosemont, IL, USA). Chicken anti-mouse IgG-Alexa 488 (A-21200) and goat anti-rabbit-Alexa 568 (A-11011) were obtained from Invitrogen (Calsbad, CA, USA). Except where indicated, all other materials are obtained from the Sigma Chemical Co. (St. Louis, MO, USA)^{17 (link)}.

Lee J.W., Thuy P.X., Koo J.H, & Moon E.Y. (2022). Nuclear factor (erythroid-derived 2)-like 2 counter-regulates thymosin beta-4 expression and primary cilium formation for HeLa cervical cancer cell survival. Scientific Reports, 12, 20170.

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Characterization of SARS-CoV-2 Spike Protein

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Ursodeoxycholic acid (UDCA, 15121) was purchased from Cayman Chemical (Ann Arbor, MI, USA). MTT [3(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide] (M5655), DAPI (D9542) and bovine serum albumin (BSA, A3059) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). CellTiter-Glo® Luminescent cell viability assay kit (G9241) was purchased from Promega Co. (Madison, WI, USA). Mouse antibodies which are reactive with acetylated tubulin (T7451) were from Sigma-Aldrich Co. (St. Louis, MO, USA). Polyethylenimine (PEI, 02371) was purchased from Polysciences, Inc. (Warrington, PA, USA). Rabbit antibodies which are reactive with Flag (14793) were from Cell Signaling Technology Inc. (Danvers, MA, USA). FITC-conjugated streptavidin (554061) and PE-conjugated streptavidin (554060) were purchased from BD Bioscience (Franklin Lakes, NJ, USA). Flag-tagged human pCMV-ACE2 plasmid (HG10108-CF) and a biotinylated SARS-CoV-2 (2019-nCoV) spike protein, RBD (40592-V08B-B) were purchased from Sino Biological Inc. (Wayne, PA, USA). Fibronectin (5050) and bovine collagen type I (5005) were purchased from Advanced BioMatrix, Carlsbad, C, USA). Except where indicated, all other materials are obtained from the Sigma Chemical Co. (St. Louis, MO, USA).

Thuy P.X., Bao T.D, & Moon E.Y. (2022). Ursodeoxycholic acid ameliorates cell migration retarded by the SARS-CoV-2 spike protein in BEAS-2B human bronchial epithelial cells. Biomedicine & Pharmacotherapy, 150, 113021.

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Western Blot Analysis of Cellular Proteins

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Cell pellets were suspended in lysis buffer (50 mmol/L Tris–HCl pH 7.4, 150 mmol/L NaCl, 1% NP40, 10 mmol/L NaF, 1 mmol/L PMSF, and protease inhibitor cocktail). Next, the lysis reaction was carried out for 15 min at 4 °C, samples were centrifuged at 13,000 rpm for 30 min at 4 °C, and protein concentration quantified by Bradford assay (Bio-Rad Protein Assay Dye Reagent Concentrate, #5000006). A total of 50 μg of each sample was loaded on 8%, 10%, or 15% polyacrylamide gels and electro-blotted on nitrocellulose membrane. Immunoreactive signals were detected with a horseradish peroxidase-conjugated secondary antibody (Bio-Rad, #1705046, #1706464). Primary antibodies were: AR (sc-52309), P53 (sc-126), tubulin (sc-5286), and SQSTM1 (sc-48402) purchased from Santa Cruz Biotechnology (Dallas, USA); BAX (ab53154), caspase 9 (ab9502), caspase 8 (ab9746), BCL2 (ab28725), p-eNOS (ab184154), eNOS (ab5589), p-AKT (ab81283) and LSD1 (ab17721) from Abcam. Fatty Acid and Lipid Metabolism Antibody Sampler Kit (#8335), GLUT1 (D3J3 A) and GAPDH (D16H11) were purchased from Cell Signaling Technology (Danvers, MA, USA); acetylated-tubulin (T7451) from Sigma–Aldrich. All antibodies were used according to the manufacturer's instructions. Semi-quantitative analysis was performed using ImageJ software (version 1.44), and the relative intensities are reported in figures.

Chianese U., Papulino C., Passaro E., Evers T.M., Babaei M., Toraldo A., De Marchi T., Niméus E., Carafa V., Nicoletti M.M., Del Gaudio N., Iaccarino N., Randazzo A., Rotili D., Mai A., Cappabianca S., Mashaghi A., Ciardiello F., Altucci L, & Benedetti R. (2022). Histone lysine demethylase inhibition reprograms prostate cancer metabolism and mechanics. Molecular Metabolism, 64, 101561.

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Dimethyloxallyl Glycine-Induced Oxidative Stress

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Dimethyloxallyl Glycine (DMOG, 71210) was from Cayman Chemical (Ann Arbor, MI, USA). N-acetyl-L-cysteine (NAC), hydrogen peroxide (H₂O₂), MTT [3(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide], sodium butyrate and 4′,6-diamidno-2-phenylinole (DAPI) were purchased from the Sigma Chemical Co. (St. Louis, MO, USA). 2’,7’-dichlorofluorescin diacetate (DCF-DA) was purchased from Molecular Probe (Eugene, OR, USA). Ciliobrevin A was from Selleck Chemicals (Houston, TX, USA). PD98059 was from Enzo Life Sciences, Inc. (Farmingdale, NY, USA). Mouse antibodies which are reactive with acetylated tubulin (T7451), and β-tubulin (T4026) were from Sigma-Aldrich Co. (St. Louis, MO, USA). Rabbit antibodies which are reactive with Nrf2 and HIF-1α were from Cell signaling Technology (Cat# 12721, Danvers, MA, USA) and Novus Biologicals (Centennial, CO, USA), respectively. Rabbit antibodies which are reactive with Arl13b (17711-1-AP) were from Proteintech Group Inc. (Rosemont, IL, USA). Chicken anti-mouse IgG-Alexa 488 (A-21200) and goat anti-rabbit-Alexa 568 (A-11011) were obtained from Invitrogen (Calsbad, CA, USA). Except where indicated, all other materials are obtained from the Sigma Chemical Co. (St. Louis, MO, USA).

Lee J.W., Cho J.Y., Thuy P.X, & Moon E.Y. (2022). HeLa Cervical Cancer Cells Are Maintained by Nephronophthisis 3-Associated Primary Cilium Formation via ROS-Induced ERK and HIF-1α Activation under Serum-Deprived Normoxic Condition. International Journal of Molecular Sciences, 23(23), 14500.

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