Overview

  • Product nameAnti-beta Actin antibody [AC-15]
    See all beta Actin primary antibodies
  • Description
    Mouse monoclonal [AC-15] to beta Actin
  • SpecificityThis antibody makes a good loading control antibody and we recommend its use as such. We also recommend ab8226, another excellent mouse monoclonal beta actin loading control antibody.
  • Tested applicationsICC/IF, Dot Blot, IHC-FoFr, ICC, IHC-P, IHC-Fr, Indirect ELISA, WB, ELISA, IHC-FrFlmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Sheep, Rabbit, Chicken, Guinea pig, Hamster, Cow, Cat, Dog, Human, Fish, Opossum, Marmoset (common), Meriones unguiculatus

    Does not react with

    Fruit fly (Drosophila melanogaster), Dictyostelium discoideum
  • Immunogen

    Synthetic peptide corresponding to beta Actin aa 1-14 (N terminal) conjugated to Keyhole Limpet Haemocyanin (KLH). Slightly modified β-cytoplasmic actin N-terminal peptide, Ac-Asp-Asp-Asp-Ile-Ala-Al​a-Leu-Val-Ile-Asp-Asn-Gl y​-Ser-Gly-Lys, conjugated to KLH.
    Sequence:

    DDDIAALVIDNGSGK

  • EpitopeN-terminal of the beta isoform of actin.
  • Positive control
    • Cultured human or chicken fibroblasts as described in Liao et al.
  • General notes

    Immunofluoresence staining of chicken gizzard ultra-thin sections (North et al. J. Cell Sci. 107:445-455 (1994)) labels the dense bodies, longitudinal channels and membrane associated dense-plaque.

Properties

Applications

Our Abpromise guarantee covers the use of ab6276 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/IF 1/200.
Dot Blot Use at an assay dependent concentration.
IHC-FoFr Use at an assay dependent concentration.
ICC 1/1000.
IHC-P 1/10000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
IHC-Fr Use at an assay dependent concentration.
Indirect ELISA Use at an assay dependent concentration.
WB 1/5000 - 1/16000. Predicted molecular weight: 42 kDa.
ELISA Use at an assay dependent concentration.
IHC-FrFl Use at an assay dependent concentration.
  • Application notesIs unsuitable for IP.
  • Target

    Anti-beta Actin antibody [AC-15] images



    • Predicted band size : 42 kDa

      David Grimm, Yale University, USA.

      MDCK cells induced with increasing amounts of doxycycline to control expression of the gene of interest. All cells were normalized for loading with an albumin protein standard asssay. Anti-beta actin (ab6276) was used at a concentration of 1:5000 in a milk blocking solution. B-actin blotting confirms the albumin assay in showing that an equal amount of lysate was loaded in each lane.

      MDCK cells induced with increasing amounts of doxycycline to control expression of the gene of interest. All cells were normalized for loading with an albumin protein standard asssay. Anti-beta actin (ab6276) was used at a concentration of 1:5000 in a milk blocking solution. B-actin blotting confirms the albumin assay in showing that an equal amount of lysate was loaded in each lane.

    • All lanes : Anti-beta Actin antibody [AC-15] (ab6276) at 1/5000 dilution

      Lane 1 : HeLa nuclear
      Lane 2 : HeLa whole cell lysate
      Lane 3 : A431 cell lysate
      Lane 4 : Jurkat cell lysate
      Lane 5 : HEK293 cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      Alexa Fluor anti mouse at 1/5000 dilution

      Performed under reducing conditions.

      Predicted band size : 42 kDa
      Observed band size : 42 kDa


    • Predicted band size : 42 kDa
      Observed band size : 42 kDa

      This image is courtesy of an Abreview submitted by Dr Mark Elliott

      Western Blot of ab6276 (used as loading control) with whole tissue lysate of grey matter from BA20 (temporal cortex).  Ab6276 was diluted 1/50000 and incubated with the sample for 16 hours at 4°C.  5 µg of lysate was loaded onto the gel, which was blocked with 5% milk for 1 hour at 20°C.  An Alexa Fluor® 680 conjugated goat anti-mouse antibody, diluted 1/5000, was used as the secondary.

      Bands below actin in image are synaptophysin (SYN).

      See Abreview

    • ab6276 at 1/500 staining human macrophage cells by ICC/IF. The cells were paraformaldehyde fixed and blocked with serum prior to incubation with the antibody for 1 hour. A Cy2 ® conjugated donkey antibody was used as the secondary.

      See Abreview

    • ab6276 at 1/10000 staining mouse brain tissue sections by IHC-P. The mouse brain was immersion-fixed in 4% Formalin/PBS for 2 days, then bisected mid-saggitally. The tissue was further fixed in same solution for a further 3 days. Both halves were processed to pwax on a dip-n-dunk tissue processor using a 20hr-cycle. Sections were cut at 5microns,floated out on water at 47C, mounted on Superfrost Plus slides and dried/melted for 24hrs in a 62C oven.

      The whole brain shows positivity but there are areas/cells that are enriched. Note that the axonal tracks are negatively stained at this dilution factor, yet some cells within those tracks are still strongly positive (many similar cells outside of the tracks are also positive).

      See Abreview

    • ab6276 staining beat actin in rat hypothalamus primary cells by ICC/IF. The cells were formaldehyde fixed, permeabilized in 0.5% Triton X-100 and blocked in 5% serum for 20 minutes at 25°C. The primary antibody was diluted 1/1000 in PBS (0.1% Triton X-100, 1% goat serum) and incubated with sample for 16 hours at 4°C. An Alexa Fluor® 546 conjugated goat polyclonal to mouse IgG1, diluted 1/500 was used as secondary.

      See Abreview

    • ab6276 detecting beta actin in AGS Human gastric carcinoma cells by direct ELISA. Samples were blocked with 5% BSA for 1 hour at 23°C and incubated with the primary antibody (1/1000 in blocking buffer) for 16 hours at 4°C. Ab6729 (1/1000) was used as the secondary antibody.

      See Abreview

    References for Anti-beta Actin antibody [AC-15] (ab6276)

    This product has been referenced in:
    • Adeosun SO  et al. Cognitive deficits and disruption of neurogenesis in a mouse model of apolipoprotein E4 domain interaction. J Biol Chem 289:2946-59 (2014). Mouse . Read more (PubMed: 24324264) »
    • Darbre PD Hypersensitivity and growth adaptation of oestrogen-deprived MCF-7 human breast cancer cells. Anticancer Res 34:99-105 (2014). WB ; Human . Read more (PubMed: 24403449) »

    See all 442 Publications for this product

    Product Wall

    Application Western blot
    Loading amount 30 µg
    Gel Running Conditions Reduced Denaturing (12)
    Sample Human Tissue lysate - whole (CNS tissue)
    Specification CNS tissue
    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 Aug 01 2014

    Application Immunocytochemistry/ Immunofluorescence
    Blocking step BSA as blocking agent for 45 minute(s) · Concentration: 2% · Temperature: RT°C
    Sample Human Cell (HeLa cells)
    Specification HeLa cells
    Permeabilization Yes - 0.2% Triton-X100
    Fixative Paraformaldehyde
    Username

    Abcam user community

    Verified customer

    Submitted May 23 2014

    Application Western blot
    Loading amount 75 µg
    Gel Running Conditions Reduced Denaturing (15)
    Sample Mouse Tissue lysate - whole (Brain Tissue)
    Specification Brain Tissue
    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 May 15 2014

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application Western blot
    Loading amount 25 µg
    Gel Running Conditions Reduced Denaturing (12.5%)
    Sample Mouse Cell lysate - whole cell (Mouse brain lysates)
    Specification Mouse brain lysates
    Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
    Username

    Herr Dr. Vladimir Milenkovic

    Verified customer

    Submitted Jul 10 2013

    Western blot

    Excellent
    Abreviews
    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application Western blot
    Loading amount 40 µg
    Gel Running Conditions Reduced Denaturing (12%)
    Sample Human Cell lysate - whole cell (IMR32-Human Neuroblastoma)
    Specification IMR32-Human Neuroblastoma
    Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
    Username

    Abcam user community

    Verified customer

    Submitted Jun 13 2013

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application Western blot
    Sample Meriones unguiculatus Cell lysate - whole cell (Liver)
    Loading amount 70 µg
    Specification Liver
    Gel Running Conditions Reduced Denaturing (10)
    Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Username

    Abcam user community

    Verified customer

    Submitted May 17 2013

    That's fine! I will send you the product asap. However in order to proceed with your order I would need the order number of previously purchased vial of ab6276.

    I will look forward o hearing from you soon.

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application Western blot
    Sample Human Cell lysate - whole cell (Melanoma)
    Loading amount 50 µg
    Specification Melanoma
    Gel Running Conditions Reduced Denaturing (7,5 %)
    Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 4% · Temperature: RT°C
    Username

    Abcam user community

    Verified customer

    Submitted Nov 30 2012

    Thank you for contacting us.

    We use to get ab49846 from our collaborator laboratory; they unfortunately have stopped making it, which is why we unpublished the antibody. Because ab49876 and ab82618 are of same clone so we should not expect hug...

    Read More

    Thank you very much for your email.

    I am sorry to hear that the results did not improve. The antibody should have worked with your protocol so it really seems, it was bad vial. We have unfortunately unpublished the antibody ab49846 due to prod...

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