Anti-alpha Tubulin antibody - Loading Control (ab89984)

Overview

  • Product nameAnti-alpha Tubulin antibody - Loading ControlSee all alpha Tubulin primary antibodies ...
  • Description
    Chicken polyclonal to alpha Tubulin - Loading Control
  • Tested applicationsWB, ICC/IF more details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Chicken, Cow, Monkey
  • Immunogen

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

    (Peptide available as ab23537.)

  • Positive control
    • This antibody gave a positive signal in the following whole cell lysates: HeLa; HEK293; HepG2; Caco2; HCT116; NIH 3T3; PC12;

Properties

Applications

Our Abpromise guarantee covers the use of ab89984 in the following tested applications.

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

Application Notes
WB Use a concentration of 1 µg/ml. Detects a band of approximately 53 kDa (predicted molecular weight: 50 kDa).Can be blocked with Human alpha Tubulin peptide (ab23537).
ICC/IF Use a concentration of 5 µg/ml.

Target

  • RelevanceMicrotubules are involved in a wide variety of cellular activities ranging from mitosis and transport events to cell movement and the maintenance of cell shape. Tubulin itself is a globular protein which consists of two polypeptides (alpha and beta tubulin). Alpha and beta tubulin dimers are assembled to 13 protofilaments that form a microtubule of 22 nm diameter. Tyrosine ligase adds a C-terminal tyrosin to monomeric alpha tubulin. Assembled microtubules can again be detyrosinated by a cytoskeleton associated carboxypeptidase. Detyrosinated alpha tubulin is referred to as Glu-tubulin. Another post-translational modification of detyrosinated alpha tubulin is C-terminal polyglutamylation which is characteristic for microtubules in neuronal cells and the mitotic spindle. Alpha tubulin is not suitable as a loading control in adipose tissue as expression of tubulin in adipose tissue is very low ( Spiegelman and Farmer, Cell, 1982, 29(1): 53-60, "in cells undergoing adipose differentiation actin synthesis decreases by 90%").
  • Cellular localizationMajor constituent of Microtubules
  • Database links
  • Alternative names
    • TUBA1 antibody
    • TUBA1A antibody
    • TUBA2 antibody
    • TUBA3 antibody
    • TUBA3C antibody
    • TUBA4A antibody
    • Tubulin, alpha 1a antibody
    • Tubulin, alpha 3c antibody
    • Tubulin, alpha 4a antibody
    see all

Anti-alpha Tubulin antibody - Loading Control images

  • All lanes : Anti-alpha Tubulin antibody - Loading Control (ab89984) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate
    Lane 3 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
    Lane 4 : Caco 2 (Human colonic carcinoma cell line) Whole Cell Lysate
    Lane 5 : HCT 116 (Human Colorectal Carcinoma) Whole Cell Lysate
    Lane 6 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Chicken IgY H&L (Alexa Fluor® 790) (ab175787) at 1/10000 dilution

    Predicted band size : 50 kDa
    Observed band size : 52 kDa (why is the actual band size different from the predicted?)

    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab89984 overnight at 4°C. Antibody binding was detected using ab175787 at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

  • All lanes : Anti-alpha Tubulin antibody - Loading Control (ab89984) at 1 µg/ml

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

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat polyclonal to Chicken IgY - H&L (HRP) at 1/3000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 50 kDa
    Observed band size : 53 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 125 kDa,37 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 30 seconds
  • ICC/IF image of ab89984 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 ab89984 at 5ug overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 Goat anti- chicken IgY (H+L) used at a 1/1000 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. This antibody also gave a positive result in HepG2 PFA fixed cell types at 5ug/ml.


  • developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 50 kDa


    Exposure time : 30 seconds

    Western blot image using the Optiblot Reducing Electrophoresis Kit - 10 x 10 cm (4-20%) (ab119220) with the
    Prism Ultra Protein Ladder (ab116028) 5µl used. We recommend using our ECL substrate kit (ab65623) .
    20ug of Lysate per lane and detection using ab89984 diluted to 1ug/ml.
    Lane 1: Hela cell lysate
    Lane 2: Jurkat cell lysate
    Lane 3: A431 cell lysate
    Lane 4: HEK293 cell lysate
    Lane 5: HepG2 cell lysate

References for Anti-alpha Tubulin antibody - Loading Control (ab89984)

This product has been referenced in:
  • Fan J  et al. Crim1 maintains retinal vascular stability during development by regulating endothelial cell Vegfa autocrine signaling. Development 141:448-59 (2014). WB ; Human . Read more (PubMed: 24353059) »
  • Bose AK & Janes KA A high-throughput assay for phosphoprotein-specific phosphatase activity in cellular extracts. Mol Cell Proteomics 12:797-806 (2013). WB . Read more (PubMed: 23233447) »

See all 4 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Sample Fruit fly (Drosophila melanogaster) Cell (Drosophila S2 cells)
Specification Drosophila S2 cells
Fixative Formaldehyde
Permeabilization Yes - Triton X-100
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Username

Ms. Zuni Bassi

Verified customer

Submitted May 20 2011

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"