Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-beta III Tubulin antibody [EP1569Y] - BSA and Azide free (ab221935)

Overview

  • Product name

    Anti-beta III Tubulin antibody [EP1569Y] - BSA and Azide free
    See all beta III Tubulin primary antibodies
  • Description

    Rabbit monoclonal [EP1569Y] to beta III Tubulin - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: IHC-P, IP, Flow Cyt, IHC-Fr, ICC/IF, WBmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human, Zebrafish
  • Immunogen

    Synthetic peptide within Human beta III Tubulin aa 400 to the C-terminus (C terminal). The exact sequence is proprietary.
    Database link: Q13509

  • Positive control

    • This antibody gave a positive signal within WB in PC12 whole cell lysate as well as the following tissue lysates: Mouse Brain; Rat Brain; Mouse Spinal Cord; Rat Spinal Cord. HeLa cytoplasmic lysate and Human breast carcinoma tissue ICC/IF: NGF-differentiated PC12 cells Flow Cyt: U-87MG and HeLa cells.
  • General notes

    Ab221935 is the carrier-free version of ab52623. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab221935 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.??

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Constituent: PBS
  • Concentration information loading...
  • Purity

    Affinity purified
  • Clonality

    Monoclonal
  • Clone number

    EP1569Y
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab221935 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-P Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IHC-Fr Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 55 kDa.

Target

  • Function

    Tubulin is the major constituent of microtubules. It binds two moles of GTP, one at an exchangeable site on the beta chain and one at a non-exchangeable site on the alpha-chain. TUBB3 plays a critical role in proper axon guidance and mantainance.
  • Tissue specificity

    Expression is primarily restricted to central and peripheral nervous system.
  • Involvement in disease

    Defects in TUBB3 are the cause of congenital fibrosis of extraocular muscles type 3A (CFEOM3A) [MIM:600638]. A congenital ocular motility disorder marked by restrictive ophthalmoplegia affecting extraocular muscles innervated by the oculomotor and/or trochlear nerves. It is clinically characterized by anchoring of the eyes in downward gaze, ptosis, and backward tilt of the head. Congenital fibrosis of extraocular muscles type 3 presents as a non-progressive, autosomal dominant disorder with variable expression. Patients may be bilaterally or unilaterally affected, and their oculo-motility defects range from complete ophthalmoplegia (with the eyes fixed in a hypo- and exotropic position), to mild asymptomatic restrictions of ocular movement. Ptosis, refractive error, amblyopia, and compensatory head positions are associated with the more severe forms of the disorder. In some cases the ocular phenotype is accompanied by additional features including developmental delay, corpus callosum agenesis, basal ganglia dysmorphism, facial weakness, polyneuropathy.
  • Sequence similarities

    Belongs to the tubulin family.
  • Domain

    The highly acidic C-terminal region may bind cations such as calcium.
  • Post-translational
    modifications

    Some glutamate residues at the C-terminus are polyglutamylated. This modification occurs exclusively on glutamate residues and results in polyglutamate chains on the gamma-carboxyl group. Also monoglycylated but not polyglycylated due to the absence of functional TTLL10 in human. Monoglycylation is mainly limited to tubulin incorporated into axonemes (cilia and flagella) whereas glutamylation is prevalent in neuronal cells, centrioles, axonemes, and the mitotic spindle. Both modifications can coexist on the same protein on adjacent residues, and lowering glycylation levels increases polyglutamylation, and reciprocally. The precise function of such modifications is still unclear but they regulate the assembly and dynamics of axonemal microtubules.
  • Cellular localization

    Cytoplasm > cytoskeleton.
  • Information by UniProt
  • Database links

  • Alternative names

    • beta 3 tubulin antibody
    • beta 4 antibody
    • beta-4 antibody
    • CDCBM antibody
    • CDCBM1 antibody
    • CFEOM3 antibody
    • CFEOM3A antibody
    • FEOM3 antibody
    • M(beta)3 antibody
    • M(beta)6 antibody
    • MC1R antibody
    • Neuron specific beta III Tubulin antibody
    • Neuron-specific class III beta-tubulin antibody
    • QccE-11995 antibody
    • QccE-15186 antibody
    • TBB3_HUMAN antibody
    • Tubb 3 antibody
    • TUBB3 antibody
    • TUBB4 antibody
    • Tubulin beta 3 antibody
    • Tubulin beta 3 chain antibody
    • Tubulin beta 4 antibody
    • Tubulin beta III antibody
    • Tubulin beta-3 chain antibody
    • Tubulin beta-4 chain antibody
    • Tubulin beta-III antibody
    • tuj 1 antibody
    • tuj1 antibody
    see all

Images

  • This WB data was generated using the same anti-beta III tubulin antibody clone, EP1569Y, in a different buffer formulation (cat# ab52623).

    Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: beta III Tubulin knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20 µg)
    Lane 4: HEK293 whole cell lysate (20 µg)

     

    Lanes 1 - 4: Merged signal (red and green). Green - ab52623 observed at 52 kDa. Red - loading control, ab8245, observed at 37 kDa.

     

    Ab52623 was shown to specifically react with beta III Tubulin in wild-type cells as signal was lost in beta III Tubulin knockout HAP1 cells. Wild-type and beta III Tubulin knockout samples were subjected to SDS-PAGE. Ab52623 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling beta III Tubulin with ab52623 at 1/20 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52623).

  • Overlay histogram showing U-87MG cells stained with ab52623 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab52623, 1/1000 dilution) for 30 min at 22ºC. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150081) at 1/2000 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (ab172730, 0.1μg/1x106) used under the same conditions. Unlabelled sample (blue line) was also used as a control.

    Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

    This antibody gave a positive signal in U-87MG cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52623).

  • ab52623 staining beta III Tubulin in NGF-differentiated PC12 cells. The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab52623 at 5μg/ml and ab7291 at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an AlexaFluor®488 Goat anti-Rabbit secondary (ab150081) at 2 μg/ml (shown in green) and AlexaFluor®594 Goat anti-Mouse secondary (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.
    Negative controls: 1– Rabbit primary and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52623).

  • ab52623, at a 1/50 dilution, staining human class III beta Tubulin in breast carcinoma tissue using Immunohistochemistry, Paraffin embedded tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52623).

  • This ICC/IF data was generated using the same anti-beta III tubulin antibody clone, EP1569Y, in a different buffer formulation (cat# ab52623).

    ab52623 staining beta III Tubulin in wild-type HAP1 cells (top panel) and TUBB3 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab52623 at 1/500 dilution and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • This IHC-Fr data was generated using the same anti-beta III tubulin antibody clone, EP1569Y, in a different buffer formulation (cat# ab52623).

    IHC-Fr image of Beta III Tubulin staining on zebrafish retina sections using ab52623 (1:100). The sections were paraformaldehyde and permeabilized using Triton-X. Antigen retrieval was performed using Tris-HCl and blocking was perfomed using 5% BSA for 1 hour at 23°C. ab38738 was diluted 1:100 and incubated with the sections for 16 hours at 4°C. The secondary antibody was goat polyclonal to rabbit IgG conjugated to Alexa Fluor 488 (1:1000).

References

This product has been referenced in:

  • Du J  et al. Overexpression of Class III ß-tubulin, Sox2, and nuclear Survivin is predictive of taxane resistance in patients with stage III ovarian epithelial cancer. BMC Cancer 15:536 (2015). IHC ; Human . Read more (PubMed: 26198101) »
  • da Silva TF  et al. Peripheral nervous system plasmalogens regulate Schwann cell differentiation and myelination. J Clin Invest 124:2560-70 (2014). Mouse . Read more (PubMed: 24762439) »
See all 6 Publications for this product

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