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Anti α tubulin t5168

Manufactured by Merck Group
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Anti-α-tubulin (T5168) is a primary antibody that specifically binds to the α-tubulin subunit of microtubules. It is a useful tool for the detection and analysis of microtubules in various applications, such as immunofluorescence microscopy and Western blotting.

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Lab products found in correlation

Anti gfp antibody 3h9, by Proteintech (3 mentions) Glass bead, by Merck Group (2 mentions) Bleomycin, by Merck Group (2 mentions) Sds page loading buffer, by Thermo Fisher Scientific (2 mentions) Anti flag, by Merck Group (2 mentions) Camptothecin, by Merck Group (2 mentions) Hybond c membrane, by Thermo Fisher Scientific (2 mentions) Laemmli buffer, by Bio-Rad (2 mentions) Vivaspin concentrators, by Merck Group (2 mentions) Rhodamine phalloidin, by Thermo Fisher Scientific (2 mentions) Anti β actin a5441, by Merck Group (2 mentions) Anti cdc2, by Santa Cruz Biotechnology (1 mentions) Protease inhibitor, by Thermo Fisher Scientific (1 mentions) Anti ku80 c48e7, by Cell Signaling Technology (1 mentions) Ecl revelblot plus, by Ozyme (1 mentions) Ripa buffer, by iNtRON Biotechnology (1 mentions) Anti flag antibody f1804, by Merck Group (1 mentions) Bca assay, by Thermo Fisher Scientific (1 mentions) Iq sybr green supermix, by Bio-Rad (1 mentions) Nem e3876, by Merck Group (1 mentions)

Spelling variants of this product

α-tubulin (1242 citations) Anti-α-tubulin (920 citations) Anti-tubulin (542 citations) T5168 (212 citations) α-tubulin (T5168) (22 citations) Anti-Tubulin (T5168) (18 citations) Mouse anti-α-tubulin (T5168, (13 citations) Anti-α-tubulin antibody (T5168; (6 citations) Mouse monoclonal anti-α-tubulin (T5168) (3 citations)

32 protocols using anti α tubulin t5168

1

Cellular Stress Response Protein Analysis

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Cellular lysates were prepared from exponentially growing cell cultures treated with 40μM camptothecin (Sigma) or 5μg/ml bleomycin (Merck). Denatured cell lysates were prepared by TCA precipitation. Cells were suspended in 20% TCA and lysed mechanically using glass beads (Sigma). Following centrifugation, the TCA precipitate was suspended in SDS-PAGE loading buffer (Invitrogen) containing Tris-base. Protein extracts were directly resolved on Tris-acetate 3–8% polyacrylamide NuPAGE gels (Invitrogen). Proteins were transferred to a nitrocellulose Hybond-C membrane (Invitrogen). The membrane was blocked in PBS-T milk 5% and probed by using anti-Flag (Sigma F1804) antibody (1:5,000 dilution), anti Cdc2 (Santa-Cruz sc-53) antibody (1,1000 dilution), anti-tubulin alpha T5168 (Sigma).
Giaccherini C., Scaglione S., Coulon S., Dehé P.M, & Gaillard P.H. (2022). Regulation of Mus81-Eme1 structure-specific endonuclease by Eme1 SUMO-binding and Rad3ATR kinase is essential in the absence of Rqh1BLM helicase. PLoS Genetics, 18(4), e1010165.
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2

Cellular Lysate Preparation and Protein Analysis

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Cellular lysates were prepared from exponentially growing cell cultures treated with 40μM camptothecin (Sigma) or 5μg/ml bleomycin (Merck). Denatured cell lysates were prepared by TCA precipitation. Cells were suspended in 20% TCA and lysed mechanically using glass beads (Sigma). Following centrifugation, the TCA precipitate was suspended in SDS-PAGE loading buffer (Invitrogen) containing Tris-base. Protein extracts were directly resolved on Tris-acetate 3-8% polyacrylamide NuPAGE gels (Invitrogen). Proteins were transferred to a nitrocellulose Hybond-C membrane (Invitrogen). The membrane was blocked in PBS-T milk 5% and probed by using anti-Flag (Sigma F1804) antibody (1:5,000 dilution), anti Cdc2 (Santa-Cruz sc-53) antibody (1:1000 dilution), anti-tubulin alpha T5168 (Sigma).
Giaccherini C., Scaglione S., Coulon S., Dehé P., & Gaillard P.L. (2021). Regulation of Mus81-Eme1 structure-specific endonuclease by Eme1 SUMO-binding and Rad3ATR kinase is essential in the absence of Rqh1BLM helicase.
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3

Characterization of p53 Isoforms

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Total protein lysate was extracted from BMMCs using RIPA buffer (Intron Biotechnology, Korea) containing a cocktail of 1% phosphatase- and 1% protease inhibitors (Thermo Fischer Scientific, USA). Sodium dodecyl sulphate-polyacrylamide gel electrophoresis and electroblotting were performed using anti-p53 (9282, Cell Signaling Technology, US and DO-7 sc-47,698 Santa Cruz Biotechnology, US), anti-MDM2 (MABE281, Merck Millipore, US), and anti-p21WAF-1 (OP64, Merck Millipore, US) antibodies. Equal protein loading was assessed using anti-alpha tubulin (T5168, Sigma Aldrich, US) and anti-Ku80 (C48E7, Cell Signaling Technology, US) antibodies. All antibodies were diluted in 1% (w/v) non-fat milk powder in TBS–Tween. Proteins were visualized using enhanced chemiluminescence (ECL RevelBlot Plus or Intense, Ozyme, France) depending on the intensity of the signal. Protein lysate isolated from Saos-2 p53-null cell line engineered to over-express specific p53 isoforms cDNA were used as reference for p53 isoforms identification.
Oh L., Hainaut P., Blanchet S, & Ariffin H. (2020). Expression of p53 N-terminal isoforms in B-cell precursor acute lymphoblastic leukemia and its correlation with clinicopathological profiles. BMC Cancer, 20, 110.
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4

Subcellular fractionation and protein detection

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Young adult animals grown at 20°C were harvested, washed, and lysed in extraction buffer (50 mM Tris, pH7.2, 250 mM sucrose, 2 mM EDTA). Supernatants were centrifuged for 60 min at 100,000xg at 4°C. The supernatant fraction was concentrated using Vivaspin Concentrators (Sigma). The pellet fraction was resuspended in 1x Laemmli buffer (Biorad). Proteins in both fractions were separated by SDS-PAGE and blotted onto PVDF membranes. Proteins were detected using an anti-GFP antibody (3H9, Chromotek) or anti-alpha tubulin (T5168, Sigma).
Raj D., Podraza-Farhanieh A., Gallego P., Kao G, & Naredi P. (2022). Identification of C. elegans ASNA-1 domains and tissue requirements that differentially influence platinum sensitivity and growth control. PLOS Genetics, 18(12), e1010538.
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5

Subcellular fractionation and protein detection

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Young adult animals grown at 20 °C were harvested, washed and lysed in extraction buffer (50 mM Tris, pH7.2, 250 mM sucrose, 2 mM EDTA). Supernatants were centrifuged for 60 min at 100,000×g at 4 °C. The supernatant fraction was concentrated using Vivaspin Concentrators (Sigma). The pellet fraction was resuspended in 1× Laemmli buffer (Biorad). Proteins in both fractions were separated by SDS-PAGE and blotted onto PVDF membranes. Proteins were detected using anti-GFP antibody [3H9] (Chromotek) or anti-alpha tubulin [T5168] (Sigma).
Raj D., Billing O., Podraza-Farhanieh A., Kraish B., Hemmingsson O., Kao G, & Naredi P. (2021). Alternative redox forms of ASNA-1 separate insulin signaling from tail-anchored protein targeting and cisplatin resistance in C. elegans. Scientific Reports, 11, 8678.
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6

Western Blot Analysis of ASNA-1 Protein

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Synchronized young adult worms were homogenized and protein concentration determined using the BCA assay (Thermo Scientific). Samples were boiled for 10 min in reducing loading buffer (SDS/β-mercaptoethanol). Proteins were separated by SDS-PAGE and blotted onto PVDF membranes Non-reducing SDS-PAGE: Lysates were boiled for 10 min in non-reducing (without β-mercaptoethanol) loading buffer and cooled to room temperature for 10 min. To protect free cysteine thiols from post-lysis oxidation, iodoacetamide was added to the samples at a final concentration of 25 mM followed by a 30 min incubation in darkness at room temperature. Proteins were separated by SDS-PAGE and blotted onto nitrocellulose membranes. Antibodies: anti-ASNA-1 antibody8 (link), anti-GFP antibody [3H9] (Chromotek) and anti-Flag antibody [F1804] (Sigma) were used. To assess equal loading, membranes were stripped and probed with anti-alpha tubulin (T5168, Sigma). Band quantification was performed using ImageJ software65 (link).
Raj D., Billing O., Podraza-Farhanieh A., Kraish B., Hemmingsson O., Kao G, & Naredi P. (2021). Alternative redox forms of ASNA-1 separate insulin signaling from tail-anchored protein targeting and cisplatin resistance in C. elegans. Scientific Reports, 11, 8678.
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7

Comprehensive Antibody and Reagent Catalog for Research

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The following antibodies and chemicals were acquired: Anti-p16 (orb228122; Biorbyt, San Francisco, CA), anti-MKRN1 (GTX106459; Genetex, Irvine, CA), anti-p90RSK (MAB 2056; R&D Systems, Minneapolis, MN), and anti-α-tubulin (T5168; Sigma-Aldrich, St. Louis, MO) were purchased. Antibodies against TERF2IP (5433) and p21 (2947) were purchased from Cell Signaling Technology (Beverly, MA). Anti-TERT (ab32020) and anti-TERF2 (ab108997) were from Abcam (Cambridge, MA). Protease inhibitor cocktail (p8340), PMSF (36978), and NEM (E3876) were from Sigma-Aldrich.
The iQSYBR Green Supermix (1708882) was from Bio-Rad (Hercules, CA); the TL PNA kit/FITC flow cytometry kit (K5327) was from Agilent Technology (Santa Clara, CA); the FITC Annexin V apoptosis detection kit (556547) was from BD Biosciences (Franklin Lakes, NJ). We also purchased TaqMan reverse transcription reagents to generate cDNA (N808–0234; Applied Biosystems, Foster City, CA), RNeasy Plus Mini Kit (#74136; QIAGEN), a dual luciferase assay kit (E1910; Promega Life Sciences, Fitchburg, WI), and Lipofectamine 2000 transfection reagent (11668027; ThermoFisher Scientific, Waltham, MA).
Kotla S., Le N.T., Vu H.T., Ko K.A., Gi Y.J., Thomas T.N., Giancursio C., Lusis A.J., Cooke J.P., Fujiwara K, & Abe J.I. (2019). Endothelial senescence-associated secretory phenotype (SASP) is regulated by Makorin-1 ubiquitin E3 ligase. Metabolism: clinical and experimental, 100, 153962.
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8

Antibody Characterization and Validation

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All chemicals were obtained from Sigma (St. Louis, MO, USA) or Merck (Darmstadt, Germany). Antibodies were as follows: anti‐α‐tubulin (T5168) and anti‐destrin (D8815) from Sigma; anti‐p65 (8242P), anti‐MMP2 (D8N9Y, 13132), and anti‐MMP9 (G657, 2270) from Cell Signaling (Danvers, MA, USA); and anti‐p53, kindly provided by Bořivoj Vojtěšek (Masaryk Memorial Cancer Institute, Czech Republic). Anti‐mouse and anti‐rabbit antibodies were obtained from Sigma and Millipore (Burlington, MA, USA), respectively.
Pampalakis G., Zingkou E., Sidiropoulos K.G., Diamandis E.P., Zoumpourlis V., Yousef G.M, & Sotiropoulou G. (2019). Biochemical pathways mediated by KLK6 protease in breast cancer. Molecular Oncology, 13(11), 2329-2343.
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9

Immunoblotting of BMDM Lysates

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BMDCs (or bone marrow-derived macrophages, which were generated as described above for BMDCs except macrophage colony-stimulating factor (M-CSF) was used) were lysed in protease inhibitor- and phosphatase inhibitor-containing CelLytic M buffer (Sigma, Burlington, MA, USA) and 20 μg of the cell lysates were separated via 15% or 7.5% SDS-PAGE, transferred to a polyvinylidene fluoride (PVDF) membrane, stained with primary antibodies, incubated with horseradish peroxidase (HRP)-labeled Immunoglobulin G (IgG) (Bio-Rad, Hercules, CA, USA), and visualized with enhanced chemiluminescence (ECL) clarity substrate (Bio-Rad, Hercules, CA, USA). The primary antibodies were anti-MsrB1 (LF-PA0088, 1:1000 dilution, Invitrogen, Carlsbad, CA, USA), anti-STAT6 (ab32520, 1:2000 dilution, Abcam, Cambridge, UK), antiphospho-STAT6 (D8S9Y, 1:1000 dilution, Cell Signaling Technology, Danvers, MA, USA), anti-β-actin (sc-47778, 1:5000 dilution, Santa Cruz, CA, USA), and anti-α-tubulin (T5168, 1:5000 dilution, Sigma, Burlington, MA, USA).
Lee H.J., Park J.S., Yoo H.J., Lee H.M., Lee B.C, & Kim J.H. (2020). The Selenoprotein MsrB1 Instructs Dendritic Cells to Induce T-Helper 1 Immune Responses. Antioxidants, 9(10), 1021.
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10

Mitochondrial Dynamics Regulation Protocols

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Anti-DRP1 (ab56788, Abcam, Cambridge, GB) and Anti-α-Tubulin (T5168, Sigma) antibodies were used at a dilution of 1/2000 for immunoblotting. anti-MFF (HPA010968, Sigma) and anti-MFF (Proteintech, Chicago, IL, 17090–1-AP) were used at 1/50 and 1/2000, respectively, for immunofluorescence staining.
Helle S.C., Feng Q., Aebersold M.J., Hirt L., Grüter R.R., Vahid A., Sirianni A., Mostowy S., Snedeker J.G., Šarić A., Idema T., Zambelli T, & Kornmann B. (2017). Mechanical force induces mitochondrial fission. eLife, 6, e30292.
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