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α tubulin

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α-tubulin is a protein found in the cytoskeleton of eukaryotic cells. It is a component of microtubules, which are involved in various cellular processes, such as cell division, intracellular transport, and cell motility.

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Lab products found in correlation

β actin, by Developmental Studies Hybridoma Bank (2 mentions) Caspase 1, by Santa Cruz Biotechnology (2 mentions) Caspase 3 cleavage product, by Cell Signaling Technology (2 mentions) Anti b23 clone fc82291, by Merck Group (2 mentions) Myoglobin, by Agilent Technologies (2 mentions) Anti myogenin f5d, by Developmental Studies Hybridoma Bank (2 mentions) Caspase 1, by Cell Signaling Technology (2 mentions) Cy3 conjugated secondary antibody, by Jackson ImmunoResearch (1 mentions) Pullulan, by Tokyo Chemical Industry (1 mentions) Ubiquitin, by Santa Cruz Biotechnology (1 mentions) Phospho camkii, by Cell Signaling Technology (1 mentions) Gapdh, by Aviva Systems Biology (1 mentions) Verapamil hydrochloride, by Merck Group (1 mentions) Txnip, by Cell Signaling Technology (1 mentions) Camkii, by Santa Cruz Biotechnology (1 mentions) Fatty acid free bovine serum albumin, by Merck Group (1 mentions) Cleaved caspase 3, by Cell Signaling Technology (1 mentions) Palmitate, by Merck Group (1 mentions) P creb, by Cell Signaling Technology (1 mentions) P p38 mapk, by Cell Signaling Technology (1 mentions)

Close spelling variants of this product

Anti-α-tubulin (12 citations) Mouse anti-α-tubulin (7 citations) Anti-α-tubulin (12G10, (4 citations) Anti-α-tubulin primary antibody (3 citations) Anti-α-tubulin antibody (12G10) (2 citations) Anti-α-tubulin (AA4.3) (2 citations) Mouse anti-α-tubulin (12G10; (2 citations) 12G10 anti-α-tubulin monoclonal antibody (2 citations)

19 protocols using α tubulin

1

Protein Extraction and Analysis from C. elegans

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To lyse the animals roughly 1 ml of lightly packed synchronized young adult animals were frozen into pellets by dropping them into liquid nitrogen, and then ground into a fine powder with a mortar and pestle. The fine powder was immediately added to 200 uls of RIPA buffer (10mM Tris/HCI pH 7.5, 150mM NaCI, 0.5mM EDTA, 0.1% SDS, 1% Triton X-100, 0.5% Deocycholate, 1mM PMSF, and 1X HALT protease) and spun to separate out the supernatant. To extract proteins the supernatant was boiled for 10 minutes in the presence of 5% 2-mercaptoethanol and 1X SDS sample buffer. Samples were run on 4-15% polyacrylamide gradient gel, and blotted onto a PVDF membrane. Antibodies and dilutions used for detecting tagged PRG-1 and tubulin were α-HA (Sigma-Aldrich, 1:5000) and α-tubulin (E7; Developmental Studies Hybridoma Bank, 1:250). Cy3-conjugated secondary antibodies (Jackson Laboratory) and a Typhoon Scanner were used for detection.
Wahba L., Hansen L, & Fire A.Z. (2021). An essential role for the piRNA pathway in regulating the ribosomal RNA pool in C. elegans. Developmental cell, 56(16), 2295-2312.e6.
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2

Pullulan-based Fluorescent Probe Synthesis

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Pullulan was purchased from the Tokyo Chemical Industry (Tokyo, Japan). Isopropyl alcohol, citric acid, fluorescein isothiocyanate isomer I (FITC), sodium hydroxide (NaOH, 99.0%), monochloroacetic acid (ClCH2COOH), lithium aluminum hydride (LiAlH4), phosphate buffered saline (PBS, pH 7.4), and dialysis tubing cellulose membranes (Mw. cutoff = 14 K), (±)-verapamil hydrochloride, fatty acid-free bovine serum albumin (BSA), and palmitate were purchased from Sigma Aldrich (St. Louis, MO, USA). Glycerol, methanol, and ethanol were purchased from Duksan Chemicals (Seoul, South Korea). Immunoblotting and immunostaining were performed using antibodies against p62, NLRP3, cleaved caspase-3, phospho-CaMKII, TXNIP (Cell Signaling Technology, Danvers, MA, USA), p62 (Sigma-Aldrich), ubiquitin, CaMKII, caspase-1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA), α-tubulin, β-actin (Developmental Studies Hybridoma Bank, Iowa City, IA, USA), and GAPDH (Aviva Systems Biology, San Diego, CA, USA). Triple-distilled and deionized water were used throughout the experiment.
Kim D.K., Han D., Bae J., Kim H., Lee S., Kim J.S., Jeong Y.G., Shin J, & Park H.W. (2023). Verapamil-loaded supramolecular hydrogel patch attenuates metabolic dysfunction-associated fatty liver disease via restoration of autophagic clearance of aggregated proteins and inhibition of NLRP3. Biomaterials Research, 27, 4.
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3

Western Blot Analysis of Cellular Proteins

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Tissue or cell lysates were prepared in RIPA buffer as previously describe46 (link). Equal protein amounts were to SDS-PAGE electrophoresis. Separated proteins on gels were then transferred onto PVDF membranes, blocked in 5% skim milk for 1 h before being incubated overnight at 4 °C. In the following primary antibodies: UCP1 (Abcam, ab10983, 1:10,000), p-p38 MAPK (Cell Signaling, 4511, 1:1000), p-ERK (Cell Signaling, 9101, 1:1000), ERK (Cell Signaling, 9102, 1:1000), p-CREB (Cell Signaling, 9196, 1:1000), GFP (Sigma-Aldrich, G1544, 1:3000) and α-tubulin (Developmental Studies Hybridoma Bank, 12G10, 1:5000). Membranes were washed with 1X TBST and then were incubated with anti-rabbit IgG (Invitrogen, 31460, 1:3000) or anti-mouse IgG (Invitrogen, 62-6520, 1:2000). Blots were visualized by ImageQuant™ LAS 4000 or X-ray film. Blots densitometry were measured using ImageJ software.
Kim S.K., Tran L.T., NamKoong C., Choi H.J., Chun H.J., Lee Y.H., Cheon M., Chung C., Hwang J., Lim H.H., Shin D.M., Choi Y.H, & Kim K.W. (2023). Mitochondria-derived peptide SHLP2 regulates energy homeostasis through the activation of hypothalamic neurons. Nature Communications, 14, 4321.
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4

Western Blotting Protein Detection Protocol

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Total cell extracts were prepared 52 (link) and western blotting was performed using antibodies against HuR (3A2, 1:10,000) 53 (link), NPM (anti-B23 clone FC82291, Sigma-Aldrich,1:30000), My-HC (Developmental studies Hybridoma5 (link),1:1000), Myoglobin (DAKO,1:500), α–tubulin (Developmental studies Hybridoma Bank, 1:1000), GFP (Clontech, Mountain View, CA, USA, 1:1000), and KSRP (affinity-purified rabbit serum 17 (link), 1:3000), anti-myogenin (F5D, obtained from Developmental Studies Hybridoma Bank, 1:250) and caspase 3 cleavage product (Cell Signaling Technology, 1:1000).
Cammas A., Sanchez B.J., Lian X.J., Dormoy-Raclet V., van der Giessen K., de Silanes I.L., Ma J., Wilusz C., Richardson J., Gorospe M., Millevoi S., Giovarelli M., Gherzi R., Di Marco S, & Gallouzi I.E. (2014). Destabilization of Nucleophosmin mRNA by the HuR/KSRP complex is required for muscle fiber formation. Nature communications, 5, 4190.
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5

Western Blotting Protein Detection Protocol

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Total cell extracts were prepared 52 (link) and western blotting was performed using antibodies against HuR (3A2, 1:10,000) 53 (link), NPM (anti-B23 clone FC82291, Sigma-Aldrich,1:30000), My-HC (Developmental studies Hybridoma5 (link),1:1000), Myoglobin (DAKO,1:500), α–tubulin (Developmental studies Hybridoma Bank, 1:1000), GFP (Clontech, Mountain View, CA, USA, 1:1000), and KSRP (affinity-purified rabbit serum 17 (link), 1:3000), anti-myogenin (F5D, obtained from Developmental Studies Hybridoma Bank, 1:250) and caspase 3 cleavage product (Cell Signaling Technology, 1:1000).
Cammas A., Sanchez B.J., Lian X.J., Dormoy-Raclet V., van der Giessen K., de Silanes I.L., Ma J., Wilusz C., Richardson J., Gorospe M., Millevoi S., Giovarelli M., Gherzi R., Di Marco S, & Gallouzi I.E. (2014). Destabilization of Nucleophosmin mRNA by the HuR/KSRP complex is required for muscle fiber formation. Nature communications, 5, 4190.
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6

Western Blot Analysis of Signaling Proteins

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IMC micromasses were lysed in SDS sample buffer (0.1% glycerol, 0.01% SDS, 0.1 M Tris, pH 6.8) on ice. Total protein concentrations were determined using the Bio-Rad DC assay (Bio-Rad). Proteins (20 μg) were separated by SDS-PAGE and transferred to nitrocellulose membranes. Membranes were blotted using antibodies for Hdac3 (Abcam, ab7030, 1:10,000 dilution), p-Thr202/Tyr204 Erk1/2 (Cell Signaling, #9101S, 1:2000 dilution), total Erk (Cell Signaling, #9102, 1:5000 dilution), p-Ser217/221 MEK (Cell Signaling, #9121, 1:5000 dilution), α-tubulin (Developmental Studies Hybridoma Bank, E7, 1:5000 dilution), β-actin (Sigma Aldrich, A5316, 1:10,000), Mmp13 (Abcam, ab39012, 1:2000), p-Ser319 Runx2 (gift from Dr. Renny Franceschi (University of Michigan), 1:1000), p-MAPK Family Antibody Sampler kit (p-p38 MAPK (Thr180/Tyr182), p-SAPK/JNK (Thr183/Tyr185), p-p44/42 MAPK (Erk1/2) (Thr202/Tyr204),) (Cell Signaling, #9910S, 1:1000), Dusp6 (Abcam, ab182605, 1:5000), Dusp7 (Abcam, ab100921, 1:5000) and corresponding secondary antibodies (Santa Cruz Biotechnology). Chemiluminescent detection was performed with Pierce femto reagent (Pierce).
Carpio L.R., Bradley E.W, & Westendorf J.J. (2016). Histone Deacetylase 3 Suppresses Erk Phosphorylation and Matrix Metalloproteinase (Mmp)-13 Activity in Chondrocytes. Connective tissue research, 58(1), 27-36.
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7

C2C12 Cell Extract Analysis

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Cell extracts were prepared by incubating undifferentiated or differentiated C2C12 cells on ice for 15 min with lysis buffer (50 mM HEPES pH 7.0, 150 mM NaCl, 10% glycerol, 1% Triton, 10 mM pyrophosphate sodium, 100 mM NaF, 1 mM EGTA, 1.5 mM MgCl2, 1× protease inhibitor (Roche)). The lysed cells were then centrifuged at 12 000 rpm for 15 min at 4°C in order to remove cell debris. The extracts were then run on an SDS-PAGE gel and transferred to nitrocellulose membranes (BioRad). Finally, the samples were analyzed by western blotting with antibodies against HuR (3A2) (39 (link)), 1:10 000), YB1 (ab12148 Abcam, 1:1000), Myog (F5D, Developmental studies Hybridoma Bank, 1:250), GFP (Takara, 1:1000), or α-tubulin (Developmental studies Hybridoma Bank, 1:1000) as loading control.
Sánchez B.J., Mubaid S., Busque S., de los Santos Y.L., Ashour K., Sadek J., Lian X.J., Khattak S., Di Marco S, & Gallouzi I.E. (2023). The formation of HuR/YB1 complex is required for the stabilization of target mRNA to promote myogenesis. Nucleic Acids Research, 51(3), 1375-1392.
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8

Quantifying Mitochondrial Proteins in Muscle

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Quadriceps muscles (~50 mg) were homogenized on ice, protein was extracted, and samples were prepared for western blotting as performed previously.9 Extracted proteins (30 μg) were electrophoresed, transferred onto nitrocellulose membranes, and prepared for antibody incubations as carried out previously.9 Antibodies used were PGC1α (#AB3242) from MilliporeSigma (Burlington, MA, USA); OPA1 (#80471), Mitofusin‐2 (#9482), VDAC (#4866), cytochrome‐C (#11940) and Cox IV (#4844) from Cell Signalling Technologies (Danvers, MA, USA); and α‐Tubulin (#12G10) from Developmental Studies Hybridoma Bank (Iowa City, IA, USA). Membranes were then incubated with either anti‐rabbit IgG (H + L) DyLight 800 or anti‐mouse IgG (H + L) DyLight 680 secondary antibodies (Cell Signalling Technologies, Danvers, MA, USA), and analysed with Odyssey's Infrared Imaging System (LI‐COR Biosciences, Lincoln, NE, USA). Total proteins were normalized to the tubulin loading control.
Huot J.R., Pin F., Chatterjee R, & Bonetto A. (2022). PGC1α overexpression preserves muscle mass and function in cisplatin‐induced cachexia. Journal of Cachexia, Sarcopenia and Muscle, 13(5), 2480-2491.
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9

Subcellular Fractionation and Western Blot Analysis

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Cells were lysed with RIPA buffer. Nuclear and cytoplasmic fractions were isolated using NE-PER Nuclear and Cytoplasmic extraction reagents (Pierce). Protein concentrations were determined using the DCTM Protein Assay (Bio-Rad). Following SDS-PAGE, Western Blot analysis was performed using the following primary antibodies: β-catenin (Millipore), NFAT1 (Cell Signaling), pan-Akt (Cell Signaling), phospho-Akt (Ser473, Cell Signaling), TATA-binding protein (TBP, Abcam), and α-tubulin (Developmental Studies Hybridoma Bank). A horseradish peroxidase-conjugated antibody (Millipore) was used as a secondary antibody, and Pierece ECL Western Detection kit (ThermoFisher) was used for protein visualization.
Gibson A.L., Hui Mingalone C.K., Foote A.T., Uchimura T., Zhang M, & Zeng L. (2017). Wnt7a Inhibits IL-1β Induced Catabolic Gene Expression and Prevents Articular Cartilage Damage in Experimental Osteoarthritis. Scientific Reports, 7, 41823.
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10

Immunoblotting Analysis of Hippo Pathway

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Cells or tissues were lysed in RIPA buffer (150 mM sodium chloride, 50 mM Tris-HCl at pH 7.4, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 1 mM PMSF) with protease inhibitors (Roche). The proteins were separated on SDS–polyacrylamide gels and transferred onto PVDF membranes (Millipore). The blots were probed with antibodies against YAP (Cell Signaling, #4912), phospho-YAP (Ser112; Cell Signaling, #4911), phospho-YAP (Ser366; Cell Signaling, #13619), Lats1 (Cell Signaling, #3477), Lats2 (Cell Signaling, #13646), Mst1 (Cell Signaling, #ab51134), NF2 (Sigma, HPA003097), Mst2 (Epitomics, 1943-1), CTGF (Santa Cruz Biotechnology, sc-14939), α-tubulin (Developmental Studies Hybridoma Bank, 12G10), PAN-TEAD (Cell Signaling, #13295), MOB1 (Cell Signaling, #3863), phospho-MOB1 (Cell Signaling, #8699), YAP, and TAZ (Cell Signaling, #8418) and normalized by Actin (Millipore, MAB1501). Signals were detected and quantified by a LI-COR infrared imaging system. The antibodies used for coimmunoprecipitation were 14-3-3 (Santa Cruz Biotechnology, sc-732) and YAP (Novus Biologicals, NB110-58358).
Chen Q., Zhang N., Xie R., Wang W., Cai J., Choi K.S., David K.K., Huang B., Yabuta N., Nojima H., Anders R.A, & Pan D. (2015). Homeostatic control of Hippo signaling activity revealed by an endogenous activating mutation in YAP. Genes & Development, 29(12), 1285-1297.
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