Overview

  • Product name

    Anti-ARF6 antibody [EPR8357]
    See all ARF6 primary antibodies
  • Description

    Rabbit monoclonal [EPR8357] to ARF6
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, Flow Cyt, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    corresponding to Human ARF6 aa 100-200.

  • Positive control

    • WB: MCF7, 293T, HEK-293, HAP1, HeLa and A431 cell lysates; Rat kidney lysates. FC: HeLa cells ICC/IF: HeLa cells
  • General notes

     

     

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

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. Stable for 12 months at -20°C.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR8357
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab131261 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
WB 1/200. Predicted molecular weight: 20 kDa.

For unpurified use at 1/1000 - 1/10000 dilution.

Flow Cyt Use at an assay dependent concentration.
ICC/IF 1/250 - 1/500.

Target

  • Function

    GTP-binding protein involved in protein trafficking; regulates endocytic recycling and cytoskeleton remodeling. May modulate vesicle budding and uncoating within the Golgi apparatus. Functions as an allosteric activator of the cholera toxin catalytic subunit, an ADP-ribosyltransferase. Involved in the regulation of dendritic spine development (By similarity). Contributes to the regulation of dendritic branching and filopodia extension.
  • Sequence similarities

    Belongs to the small GTPase superfamily. Arf family.
  • Cellular localization

    Golgi apparatus. Cell membrane. Endosome membrane. Cell membrane. Cell projection > filopodium membrane. Recruited to the cell membrane in association with CYTH2 and ARL4C.
  • Information by UniProt
  • Database links

  • Alternative names

    • antibody
    • ADP ribosylation factor 6 antibody
    • ADP ribosylation factor protein 6 antibody
    • ADP-ribosylation factor 6 antibody
    • ARF6 antibody
    • ARF6_HUMAN antibody
    • DKFZp564M0264 antibody
    • Small GTP binding protein antibody
    • Small GTPase antibody
    see all

Images

  • Anti-ARF6 antibody [EPR8357] (ab131261) at 1/1000 dilution (Purified) + HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 20 kDa
    Observed band size: 18 kDa
    why is the actual band size different from the predicted?



    This antibody shows low affinity on HeLa cells and detects a strong non-specific band around 75 KD.

  • All lanes : Anti-ARF6 antibody [EPR8357] (ab131261) at 1/200 dilution (Purified)

    Lane 1 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : Mouse kidney lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 20 kDa
    Observed band size: 18 kDa why is the actual band size different from the predicted?

  • Anti-ARF6 antibody [EPR8357] (ab131261) at 1/1000 dilution (Purified) + Rat kidney lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 20 kDa
    Observed band size: 18 kDa why is the actual band size different from the predicted?

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: ARF6 knockout HAP1 cell lysate (20 µg)
    Lane 3: HEK293 cell lysate (20 µg)
    Lane 4: HeLa cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab131261 (unpurified) observed at 20 kDa. Red - loading control, ab8245, observed at 37 kDa.
    ab131261 was shown to recognize ARF6 when ARF6 knockout samples were used, along with additional cross-reactive bands. Wild-type and ARF6 knockout samples were subjected to SDS-PAGE. ab131261 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling ARF6 with purified ab131261 at 1/250 dilution (1.1 µg/ml). Cells were fixed in 100% Methanol. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 (2.5 µg/ml) dilution. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling ARF6 with purified ab131261 at 1/250 dilution(10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(ab150077)(1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black)(ab172730) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

  • Immunofluorescent staining of HeLa cells labelling ARF6 with ab131261 at 1/250 dilution.
  • All lanes : Anti-ARF6 antibody [EPR8357] (ab131261) at 1/1000 dilution ((unpurified))

    Lane 1 : MCF7 cell lysate
    Lane 2 : 293T cell lysate
    Lane 3 : A431 cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 20 kDa

References

This product has been referenced in:

  • Costa Verdera H  et al. Cellular uptake of extracellular vesicles is mediated by clathrin-independent endocytosis and macropinocytosis. J Control Release 266:100-108 (2017). WB . Read more (PubMed: 28919558) »
  • Pelletán LE  et al. ADP ribosylation factor 6 (ARF6) promotes acrosomal exocytosis by modulating lipid turnover and Rab3A activation. J Biol Chem 290:9823-41 (2015). IF . Read more (PubMed: 25713146) »
See all 3 Publications for this product

Customer reviews and Q&As

1-4 of 4 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Permeabilization
No
Specification
HeLa
Fixative
Methanol

Dr. Kirk Mcmanus

Verified customer

Submitted Dec 08 2017

Question
Answer

Merci de nous avoir contactés.

Nous sommes désolés d'apprendre que le produit que vous avez reçu ne fonctionne pas comme attendu. Je me permets de vous transférer le numéro de commande de remplacement gratuit (ab77581) de ce produit : xxxxx. Vous recevrez prochainement un mail de confirmation comprenant les détails d'expédition.

N'hésitez pas à nous contacter lors d'une prochaine occasion.

Read More

Answer

Merci pour ce supplément d’informations.

Malheureusement nous n’avons pas d’autre lot en stock pour ab131261 et je crains que si je vous envoie en remplacement le même lot, que vous obtiendrez les mêmes résultats.

Je peux vous proposer ab377581 (www.abcam.com/ab77581) qui est aussi garanti de fonctionner en ICC/IF chez l’humain. Sinon, je peux également vous faire un avoir ou un remboursement.

J’attends votre réponse en ce qui concerne la façon dont vous voulez procéder avec cette affaire.

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Answer

Je suis désolée d'apprendre que ab131261 ne vous donne pas de bons résultats en ICC.

Notre politique est d'offrir tout d'abord la meilleure assistance technique. Si nous concluons que le produit que vous avez reçu ne fonctionne pas comme mentionné sur sa fiche technique et que ce produit a été commandé sous 120 jours, nous nous ferons un plaisir de vous offrir un remplacement gratuit ou un avoir en compensation.

J'aimerais donc vous offrir quelques suggestions afin d'optimiser vos résultats.

Selon l’information sur “Human Protein Atlas” il semble que ARF6 pourrait être localisé au cytoplasme (http://www.proteinatlas.org/ENSG00000165527).

Avez-vous fait un control négative ou un control secondaire pour confirmer que le marquage est en effet non-spécifique?

Pourriez-vous également me faire savoir le numéro de lot d’ab131261 que vous avez reçu ?

J'espère que ces informations vous aideront. Si nos suggestions ne permettent pas d'améliorer vos résultats, n'hésitez pas à me recontacter pour plus d'assistance.

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