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Mouse Monoclonal to alpha-tubulin

TU-01 (IgG1)

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Background

The microtubules are intracellular dynamic polymers made up of evolutionarily conserved polymorphic alpha/beta-tubulin heterodimers and a large number of microtubule-associated proteins (MAPs). The microtubules consist of 13 protofilaments and have an outer diameter 25 nm. Microtubules have their intrinsic polarity; highly dynamic plus ends and less dynamic minus ends. Microtubules are required for vital processes in eukaryotic cells including mitosis, meiosis, maintenance of cell shape and intracellular transport. Microtubules are also necessary for movement of cells by means of flagella and cilia. In mammalian tissue culture cells microtubules have their minus ends anchored in microtubule organizing centers (MTOCs).The GTP (guanosintriphosphate) molecule is an essential for tubulin heterodimer to associate with other heterodimers to form microtubule. In vivo, microtubule dynamics vary considerably. Microtubule polymerization is reversible and a populations of microtubules in cells are on their minus ends either growing or shortening – this phenomenon is called dynamic instability of microtubules. On a practical level, microtubules can easily be stabilized by the addition of non-hydrolysable analogues of GTP (eg. GMPPCP) or more commonly by anti-cancer drugs such as Taxol. Taxol stabilizes microtubules at room temperature for many hours. Using limited proteolysis by enzymes both tubulin subunits can be divided into N-terminal and C-terminal structural domains.

The alpha-tubulin (relative molecular weight around 50 kDa) is globular protein that exists in cells as part of soluble alpha/beta-tubulin dimer or it is polymerized into microtubules. In different species it is coded by multiple tubulin genes that form tubulin classes (in human 6 genes). Expressed tubulin genes are named tubulin isotypes. Some of the tubulin isotypes are expressed ubiquitously, while some have more restricted tissue expression.
Alpha-tubulin is also subject of numerous post-translational modifications. Tubulin isotypes and their posttranslational modifications are responsible for multiple tubulin charge variants - tubulin isoforms. Heterogeneity of alpha-tubulin is concentrated in C-terminal structural domain.

Specificity

The antibody TU-01 recognizes the defined epitope (aa 65-97) on N-terminal structural domain of alpha-tubulin.

Regulatory Status

Immunogen

Fraction of tubulin purified from porcine brain by two cycles of polymerization - depolymerization.

Species Reactivity:

  • Broad species reactivity

Negative Species:

Applications:

  • Western Blotting
    Recommended dilution: 1-2 μg/ml, incubation 60 min in room temperature
    Positive control:
    HPB-ALL human peripheral blood leukemia cell line (incubation 60 min)
    Porcine brain lysate (incubation 90 min)
    Sample preparation: Resuspend approx. 50 mil. cells in 1 ml cold Lysis buffer (1% laurylmaltoside in 20 mM Tris/Cl, 100 mM NaCl pH 8.2, 50 mM NaF including Protease inhibitor Cocktail). Incubate 60 min on ice. Centrifuge to remove cell debris. Mix lysate with reducing Laemmli SDS-PAGE sample buffer.
    Application note: Reducing conditions.
  • Immunohistochemistry (paraffin sections)
    Recommended dilution: 5 μg/ml
    Positive tissue: heart
  • Immunocytochemistry
    Staining technique:
    fixed and permeabilized cells
Usage note:
Indicated dilutions are recommended starting points for use of this product. Working concentrations should be determined by the investigator.

Product Specific References

  • *Viklicky V, Draber P, Hasek J, Bartek J: Production and characterization of a monoclonal antitubulin antibody. Cell Biol Int Rep. 1982 Aug;6(8):725-31. [Abstract]
  • *Draber P, Draberova E, Zicconi D, Sellitto C, Viklicky V, Cappuccinelli P: Heterogeneity of microtubules recognized by monoclonal antibodies to alpha-tubulin. Eur J Cell Biol. 1986 Jun;41(1):82-8. [Abstract]
  • *Grimm M, Breitling F, Little M: Location of the epitope for the alpha-tubulin monoclonal antibody TU-O1. Biochim Biophys Acta. 1987 Jul 24;914(1):83-8. [Abstract]
  • *Draber P, Draberova E, Linhartova I, Viklicky V: Differences in the exposure of C- and N-terminal tubulin domains in cytoplasmic microtubules detected with domain-specific monoclonal antibodies. J Cell Sci. 1989 Mar;92 ( Pt 3):519-28. [Abstract] [Full Text]
  • *Draber P, Draberova E, Viklicky V: Immunostaining of human spermatozoa with tubulin domain-specific monoclonal antibodies. Recognition of a unique beta tubulin epitope in the sperm head. Histochemistry. 1991;95(5):519-24. [Abstract]
  • *Linhartova I, Draber P, Draberova E, Viklicky V: Immunological discrimination of beta-tubulin isoforms in developing mouse brain. Post-translational modification of non-class-III beta-tubulins. Biochem J. 1992 Dec 15;288 ( Pt 3):919-24. [Abstract] [Full Text]
  • *Nováková M, Dráberová E, Schürmann W, Czihak G, Viklický V, Dráber P: gamma-Tubulin redistribution in taxol-treated mitotic cells probed by monoclonal antibodies. Cell Motil Cytoskeleton. 1996;33(1):38-51. [Abstract]
  • *Smertenko A, Blume Y, Viklicky V, Opatrny Z, Draber P: Post-translational modifications and multiple tubulin isoforms in Nicotiana tabacum L. cells. Planta. 1997;201(3):349-58. [Abstract]
  • *Kukharskyy V, Sulimenko V, Macůrek L, Sulimenko T, Dráberová E, Dráber P: Complexes of gamma-tubulin with nonreceptor protein tyrosine kinases Src and Fyn in differentiating P19 embryonal carcinoma cells. Exp Cell Res. 2004 Aug 1;298(1):218-28. [Abstract]
  • *Lukas J, Mazna P, Valenta T, Doubravska L, Pospichalova V, Vojtechova M, Fafilek B, Ivanek R, Plachy J, Novak J, Korinek V: Dazap2 modulates transcription driven by the Wnt effector TCF-4. Nucleic Acids Res. 2009 Mar 20. [Epub ahead of print] [Abstract]
  • *Smertenko A, Blume Y, Viklický V, Dráber P: Exposure of tubulin structural domains in Nicotiana tabacum microtubules probed by monoclonal antibodies. Eur J Cell Biol. 1997 Feb;72(2):104-12. [Abstract]
  • For research use only. Not for drug, diagnostic or other use.

    Related Products

  • Mouse Monoclonal to alpha-tubulin TU-02 (IgM)
  • Mouse Monoclonal to alpha-tubulin TU-16 (IgM)
  • Mouse IgG1 Isotype Control
  • Example Data





    TU-01
    Fig. 1.


    Fig. 1. Immunofluorescence staining of 3T3 mouse embryonal fibroblast cell line using anti-alpha-tubulin (TU-01; green) and anti-Vimentin (VI-01; cat. no. 11-254-C100; red). Nucleus is stained with DAPI (blue).


    TU-01
    Fig. 2.


    Fig. 2. Immunofluorescence staining of HeLa human cervix carcinoma cell line using anti-alpha-tubulin (TU-01; red). Nucleus is stained with DAPI (blue).


    TU-01
    Fig. 3.


    Fig. 3. Immunofluorescence staining of 3T3 mouse embryonal fibroblast cell line using anti-alpha-tubulin (TU-01; green). Nucleus is stained with DAPI (blue).

    loading control
    Fig. 4.


    Fig. 4. Use of anti-alpha-tubulin antibody TU-01 as a loading control (A) in an Western blotting experiment revealing the staining pattern of various cell lysates by a newly developed monoclonal antibody (B).


    TU-01
    Fig. 5.

    Fig. 5. Immunoprecipitation of alpha-tubulin from HeLa and RAJI cell lysate by antibody TU-16 and its detection by antibody TU-01. IgM heavy chain (76-92 kDa) and IgM light chain (25-30 kDa) indicated. Mr of alpha tubulin is around 50 kDa.
    L = lysate
    IPr = immunoprecipitate (reducing conditions)
    IPn = immunoprecipitate (non-reducing conditions)





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