Anti-alpha Tubulin antibody - Loading Control (ab4074)

Overview

  • Product name
    Anti-alpha Tubulin antibody - Loading Control
    See all alpha Tubulin primary antibodies
  • Description
    Rabbit polyclonal to alpha Tubulin - Loading Control
  • Host species
    Rabbit
  • Specificity
    This antibody detects a single band in cell lines, corresponding to alpha tubulin.
  • Tested applications
    Suitable for: IHC-P, WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Cow, Human
    Predicted to work with: Pig, Xenopus laevis, Chimpanzee, Monkey, Zebrafish, Chinese hamster
  • Immunogen

    Synthetic peptide corresponding to Human alpha Tubulin aa 1-100 conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab23537, ab26869)

  • Positive control
    • This antibody gave a positive signal in HeLa cells in WB and IF.
  • General notes


    This antibody makes an excellent loading control.

Properties

Applications

Our Abpromise guarantee covers the use of ab4074 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.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 50 kDa (predicted molecular weight: 50 kDa). Can be blocked with alpha Tubulin peptide (ab23537).
ICC/IF Use a concentration of 1 µg/ml.

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.
  • Sequence similarities
    Belongs to the tubulin family.
  • 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.
    Acetylation of alpha chains at Lys-40 stabilizes microtubules and affects affinity and processivity of microtubule motors. This modification has a role in multiple cellular functions, ranging from cell motility, cell cycle progression or cell differentiation to intracellular trafficking and signaling.
  • Cellular localization
    Cytoplasm > cytoskeleton.
  • Information by UniProt
  • Database links
  • Alternative names
    • Alpha-tubulin 1 antibody
    • ALS22 antibody
    • B ALPHA 1 antibody
    • bA408E5.3 antibody
    • H2 ALPHA antibody
    • Hum a tub1 antibody
    • Hum a tub2 antibody
    • LIS3 antibody
    • MGC171407 antibody
    • MGC55332 antibody
    • TBA4A_HUMAN antibody
    • Testis-specific alpha-tubulin antibody
    • TUBA1 antibody
    • TUBA1A antibody
    • tuba1l antibody
    • Tuba4a antibody
    • Tubulin alpha 1 chain antibody
    • Tubulin alpha antibody
    • Tubulin alpha-1 chain antibody
    • tubulin alpha-1B chain antibody
    • Tubulin alpha-4A chain antibody
    • Tubulin H2-alpha antibody
    • Tubulin, alpha 1 (testis specific) antibody
    • tubulin, alpha 1, like antibody
    • Tubulin, alpha 4a antibody
    • Tubulin, alpha, testis-specific antibody
    • Tubulin, alpha-1 antibody
    see all

Images

  • All lanes : Anti-alpha Tubulin antibody - Loading Control (ab4074) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252)
    Lane 2 : NIH 3T3 (Mouse) Whole Cell Lysate (ab52956)
    Lane 3 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate (ab50957)

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) (ab65484) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size: 50 kDa

  • Immunocytochemistry of SisNPFR-C6E8 cell line. Cells were prefixed in freshly prepared 4% paraformaldehyde (PFA) in F12 K culture medium with 10% fetal bovine serum and Geneticin. Fixed cells were permeabilized in PBST (0.25% Triton X-100 in PBS) for 5 min and blocked at 4°C overnight in 5% normal goat serum. The cells were then incubated at 4°C overnight with ab4074 (1:50.) Cover slips were placed onto slides and mounted with VectaShield containing DAPI.

  • All lanes : Anti-alpha Tubulin antibody - Loading Control (ab4074) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252)
    Lane 2 : HEK293 (Human) Whole Cell Lysate (ab52256)
    Lane 3 : NIH 3T3 whole cell lysate (ab7179)
    Lane 4 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate (ab50957)
    Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252) with Human alpha Tubulin peptide (ab23537) at 1 µg/ml
    Lanes 6-7 : NIH 3T3 whole cell lysate (ab7179) with Human alpha Tubulin peptide (ab23537) at 1 µg/ml
    Lane 8 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate (ab50957) with Human alpha Tubulin peptide (ab23537) at 1 µg/ml

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) (ab65484) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size: 50 kDa

  • (Image courtesy of Human Protein Atlas).
    ab4074 staining alpha tubulin in the glandular cells of the endometrium. Paraffin embedded endometrium tissue of a 33 year-old female was incubated with ab4074 (1/25 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6. ab4074 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org

    .

  • ICC/IF image of ab4074 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab4074, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 phalloidin was used to label F-actin (red). 
  • All lanes : Anti-alpha Tubulin antibody - Loading Control (ab4074) at 1 µg/ml

    Lane 1 : Mouse 3T3 lysate
    Lane 2 : Rat liver lysate
    Lane 3 : Cow cell lysate
    Lane 4 : Chicken cell lysate
    Lane 5 : CHO lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 50 kDa
    Observed band size: 50 kDa


    Exposure time: 30 seconds


    ab4074 cross-reacts strongly with Mouse 3T3 lysate and shows weak cross-reactivity with Rat liver, Cow, Chicken and CHO lysates.

References

This product has been referenced in:
  • Deng WT  et al. Human L- and M-opsins restore M-cone function in a mouse model for human blue cone monochromacy. Mol Vis 24:17-28 (2018). Read more (PubMed: 29386880) »
  • Moore KS  et al. Csde1 binds transcripts involved in protein homeostasis and controls their expression in an erythroid cell line. Sci Rep 8:2628 (2018). WB . Read more (PubMed: 29422612) »

See all 160 Publications for this product

Customer reviews and Q&As

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 20°C
Sample
Human Cell (Ovarian Carcinoma (TOV112D))
Specification
Ovarian Carcinoma (TOV112D)
Permeabilization
Yes - 0.1% Triton-x100
Fixative
Paraformaldehyde
Username

Dr. Gajendra Katara

Verified customer

Submitted Aug 18 2014

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 20°C
Sample
Human Cell (SKOV-3 Ovarian Carcinoma)
Specification
SKOV-3 Ovarian Carcinoma
Permeabilization
Yes - 0.1% Triton-x100
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Aug 18 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (10%)
Sample
Mouse Cell lysate - whole cell (Mouse embryonic stem cells)
Specification
Mouse embryonic stem cells
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 Jul 09 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (mouse fibroblast)
Loading amount
50 µg
Specification
mouse fibroblast
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Mar 12 2013

Application
Western blot
Sample
Human Cell lysate - whole cell (HCT116 cell line)
Gel Running Conditions
Reduced Denaturing
Loading amount
10 µg
Treatment
No treatment (control A-B) and compound treated 1-5 for 48hrs
Specification
HCT116 cell line
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 Jul 04 2018

Application
Western blot
Sample
Human Cell lysate - whole cell (Prosate cancer cell line)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
10 µg
Specification
Prosate cancer cell line
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 4°C
Username

Abcam user community

Verified customer

Submitted Dec 11 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (nasopharyngeal carcinoma)
Gel Running Conditions
Reduced Denaturing
Loading amount
20 µg
Specification
nasopharyngeal carcinoma
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C
Username

Abcam user community

Verified customer

Submitted Jan 13 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (ovarian cancer cells)
Permeabilization
Yes - methanol:acetone (9:1)
Specification
ovarian cancer cells
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Fixative
Paraformaldehyde
Username

Dr. Gajendra Katara

Verified customer

Submitted Feb 15 2016

Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (Hep G2, liver)
Specification
Hep G2, liver
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 Mar 17 2015

Thank you for your answer and clarification. This is an extremely important piece of information.

The most widely used loading controls are tubulin, actin and GAPDH. However, some chemicals can alter the expression levels of these proteins so...

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