Anti-beta Tubulin antibody - Loading Control (ab6046)

Overview

  • Product name
    Anti-beta Tubulin antibody - Loading Control
    See all beta Tubulin primary antibodies
  • Description
    Rabbit polyclonal to beta Tubulin - Loading Control
  • Specificity
    This antibody detects a single clean band at 50kD representing beta Tubulin. This band is significantly reduced by using peptide blocking.
  • Tested applications
    Suitable for: ICC, IHC-Fr, WB, ICC/IF, ELISA, IHC-P, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Human, Pig, Xenopus laevis, Zebrafish, Chinese hamster
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human beta Tubulin.

    (Peptide available as ab20775.)

  • Positive control
    • HeLa Cell lysate; A431 Cell lysate; MCF7 Cell lysate; 293 Cell lysate; HeLa Cell lysate; A431 Cell lysate; MCF7 Cell lysate; 293 Cell lysate;

Properties

Applications

Our Abpromise guarantee covers the use of ab6046 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
ICC 1/200. (see Abreview)
IHC-Fr 1/200. (see Abreview)
WB 1/500. Detects a band of approximately 50 kDa (predicted molecular weight: 50 kDa).
ICC/IF 1/200. (see PMID: 16030258)
ELISA Use at an assay dependent concentration.
IHC-P Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
IP Use at an assay dependent concentration.

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.
  • Tissue specificity
    Ubiquitously expressed with highest levels in spleen, thymus and immature brain.
  • Involvement in disease
    Cortical dysplasia, complex, with other brain malformations 6
    Skin creases, congenital symmetric circumferential, 1
  • 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, resulting in polyglutamate chains on the gamma-carboxyl group (PubMed:26875866). Polyglutamylation plays a key role in microtubule severing by spastin (SPAST). SPAST preferentially recognizes and acts on microtubules decorated with short polyglutamate tails: severing activity by SPAST increases as the number of glutamates per tubulin rises from one to eight, but decreases beyond this glutamylation threshold (PubMed:26875866).
    Some glutamate residues at the C-terminus are 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). Both polyglutamylation and monoglycylation can coexist on the same protein on adjacent residues, and lowering glycylation levels increases polyglutamylation, and reciprocally. The precise function of monoglycylation is still unclear.
    Phosphorylated on Ser-172 by CDK1 during the cell cycle, from metaphase to telophase, but not in interphase. This phosphorylation inhibits tubulin incorporation into microtubules.
  • Cellular localization
    Cytoplasm, cytoskeleton.
  • Information by UniProt
  • Database links
  • Alternative names
    • Beta 4 tubulin antibody
    • Beta 5 tubulin antibody
    • beta Ib tubulin antibody
    • Beta1 tubulin antibody
    • Class I beta tubulin antibody
    • M40 antibody
    • MGC117247 antibody
    • MGC16435 antibody
    • OK/SW cl.56 antibody
    • OK/SWcl.56 antibody
    • TBB5_HUMAN antibody
    • TUBB 1 antibody
    • TUBB 2 antibody
    • TUBB 5 antibody
    • TUBB antibody
    • TUBB1 antibody
    • TUBB2 antibody
    • TUBB5 antibody
    • tubulin beta 1 chain antibody
    • Tubulin beta 2 chain antibody
    • tubulin beta 5 chain antibody
    • Tubulin beta chain antibody
    • Tubulin beta class I antibody
    • tubulin beta polypeptide antibody
    • Tubulin beta-5 chain antibody
    see all

Images


  • Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 50 kDa


    Exposure time : 10 seconds

    Lane 1 - 8 : beta Tubulin antibody - Loading Control (ab6046) at 1/500 dilution

    Lane 1 : HeLa Cell lysate at 20 ug
    Lane 2 : A431 Cell lysate at 20 ug
    Lane 3 : MCF7 Cell lysate at 20 ug
    Lane 4 : 293 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 5 : HeLa Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 6 : A431 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 7 : MCF7 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 8 : 293 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml

  • ICC/IF image of ab6046 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then incubated in 1% BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6046, 1µg/ml) overnight at +4°C. The secondary antibody (green) was ab150081 Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    The negative control (inset) is a secondary-only assay to demonstrate low non-specific binding of the secondary antibody.

    This product also gave a positive signal under the same testing conditions in HeLa cells fixed with 100% methanol (5 min).

  • ab6046 staining beta Tubulin in human stomach tissue by Immunohistochemistry (frozen sections). Tissue was fixed with acetone and then blocked with 5% serum for 1 hour at 23°C followed by incubation with the primary antibody at a 1/200 dilution for 1 hour at 23°C. An undiluted HRP-conjugated goat polyclonal was used as secondary antibody.

    See Abreview

  • ICC/IF image of ab6046 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6046, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • Adiponectin (green) was detected using adiponectin primary antibody (ab22554; 2.5 µl/mL). Beta tubulin (red) was detected using the rabbit polyclonal (ab6046) antibody. Cells were imaged by confocal microscopy, using z-stack for adipocyte-like cells.

  • Beta Tubulin was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to Tubulin and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab6046.
    Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
    Band: 50kDa: beta Tubulin.
  • The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6046, 1µg/ml and ab37266, 1µg/ml) overnight at +4°C. The secondary antibodies were ab150115 Alexa Fluor® 647 goat anti-mouse IgG (H+L) used at 2µg/ml for 1h and ab175652 Alexa Fluor® 405 goat anti-rabbit IgG (H+L) used at 2µg/ml for 1h. Nuclear Green LCS1 (ab138904) was used to stain the cell nuclei (green) at a dilution of 1/500.

     

  • IHC image of beta Tubulin staining in human liver carcinoma FFPE section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab6046, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

  • Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 50 kDa

    Lane 1 - 8 : beta Tubulin antibody - Loading Control (ab6046) at 1/500 dilution

    Lane 1 : HeLa Cell lysate at 20 ug
    Lane 2 : A431 Cell lysate at 20 ug
    Lane 3 : MCF7 Cell lysate at 20 ug
    Lane 4 : 293 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 5 : HeLa Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 6 : A431 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 7 : MCF7 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml
    Lane 8 : 293 Cell lysate at 20 ug with beta Tubulin peptide (ab20775) at 1 ug/ml

    Secondary

References

This product has been referenced in:
  • Hempel C  et al. Binding of Plasmodium falciparum to CD36 can be shielded by the glycocalyx. Malar J 16:193 (2017). WB ; Chinese hamster . Read more (PubMed: 28486940) »
  • Wang X  et al. Unique molecular profile of exosomes derived from primary human proximal tubular epithelial cells under diseased conditions. J Extracell Vesicles 6:1314073 (2017). WB ; Human . Read more (PubMed: 28473886) »

See all 348 Publications for this product

Customer reviews and Q&As

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Rat Tissue lysate - whole (brown adipose tissue)
Gel Running Conditions
Reduced Denaturing (4-20)
Loading amount
20 µg
Treatment
in vivo delivery of viral vector to overexpress leptin in the brain
Specification
brown adipose tissue
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Sep 28 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Testis, adult)
Permeabilization
Yes - 0.1% Triton X-100 in PBS
Specification
Testis, adult
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 20°C
Fixative
Paraformaldehyde
Username

Mr. Bryan Niedenberger

Verified customer

Submitted Aug 04 2016

Application
Immunocytochemistry
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C
Sample
Mouse Cultured Cells (Mouse Embryonic Fibroblasts)
Specification
Mouse Embryonic Fibroblasts
Permeabilization
Yes - 0.5% Triton-X
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted May 26 2014

Application
Immunocytochemistry
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C
Sample
Human Cultured Cells (H460)
Specification
H460
Permeabilization
Yes - 0.5% Triton-X
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted May 26 2014

Application
Western blot
Loading amount
100 µg
Gel Running Conditions
Reduced Denaturing (8)
Sample
Human Cell lysate - whole cell (MDAMB231)
Specification
MDAMB231
Blocking step
Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: RT°C
Username

Abcam user community

Verified customer

Submitted Jan 09 2014

Application
Western blot
Loading amount
15 µg
Gel Running Conditions
Reduced Denaturing (10% GEL)
Sample
Human Cell lysate - whole cell (MCF7)
Specification
MCF7
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

Submitted Dec 13 2013

Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing (10% GEL)
Sample
Mouse Cell lysate - whole cell (MEF)
Specification
MEF
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

Submitted Dec 13 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
60 µg
Gel Running Conditions
Reduced Denaturing (8%)
Sample
Rat Tissue lysate - whole (Mammary Tumor)
Specification
Mammary Tumor
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Jun 25 2013

We would recommend the use of one of the following products for a loading control with zebrafish samples: AB15224 anti alpha Tubulin (http://www.abcam.com/alpha-tubulin-antibody-ab15246.html). This product has been featured in the following publicat...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Xenopus laevis Tissue lysate - whole (Head lysates)
Loading amount
30 µg
Specification
Head lysates
Gel Running Conditions
Reduced Denaturing (12% TGX (Bio-Rad))
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

Submitted Nov 19 2012

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