Anti-ADP Ribosylation Factor [1D9] antibody (ab2806)

Overview

  • Product nameAnti-ADP Ribosylation Factor [1D9] antibody
  • Description
    Mouse monoclonal [1D9] to ADP Ribosylation Factor
  • Tested applicationsICC/IF, ICC, ELISA, IHC, WB, Flow Cyt, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Hamster, Cow, Dog, Human, Fruit fly (Drosophila melanogaster)
    Predicted to work with: Non Human Primates
  • Immunogen

    Recombinant full length protein corresponding to Human ADP Ribosylation Factor.

Properties

Applications

Our Abpromise guarantee covers the use of ab2806 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 1/500.
ICC Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
IHC 1/100.
WB 1/500.
Flow Cyt 1/20.
IP Use at an assay dependent concentration.

Target

  • RelevanceThe ADP-ribosylation factor (Arf) family comprises a group of structurally and functionally conserved 21 kDa proteins, which are members of the Ras superfamily of regulatory GTP-binding proteins. Arf is involved in intracellular protein traffic to and within the Golgi complex. Arf has a number of disparate activities including maintenance of organelle integrity, assembly of coat proteins, as a co-factor for cholera toxin and as an activator of phospholipase D. The Arf family is divided functionally into the Arf and the Arf-like (Arl) proteins. Arfs share more than 60% sequence identity, appear to be ubiquitous in eukaryotes, and are highly conserved evolutionarily.
  • Cellular localizationGolgi Apparatus
  • Database links
  • Alternative names
    • ADP ribosylation factor 1 antibody
    • ARF 1 antibody
    • ARF antibody
    • ARF1 antibody
    • ARF4 antibody
    • ARF6 antibody
    see all

Anti-ADP Ribosylation Factor [1D9] antibody images

  • Immunocytochemistry/Immunofluorescence analysis of ADP Ribosylation Factor shows staining in HeLa cells. ADP Ribosylation Factor (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were incubated without (control) or with ab2806 (1:100) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • Immunocytochemistry/Immunofluorescence analysis of ADP Ribosylation Factor shows staining in MCF-7 cells. ADP Ribosylation Factor (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were incubated without (control) or with ab2806 (1:100) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • Immunocytochemistry/Immunofluorescence analysis of ADP Ribosylation Factor shows staining in U251 cells. ADP Ribosylation Factor (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were incubated without (control) or with ab2806 (1:100) overnight at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • Flow cytometry analysis of ADP Ribosylation Factor showing positive staining in the cytoplasm of MCF-7 cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/ml, fixed with 2% paraformaldehyde, washed with PBS, and incubated with ab2806 (1:80) for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

  • Flow cytometry analysis of ADP Ribosylation Factor showing positive staining in the cytoplasm of Hela cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/ml, fixed with 2% paraformaldehyde, washed with PBS, and incubated with ab2806 (1:80) for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

  • Flow cytometry analysis of ADP Ribosylation Factor showing positive staining in the cytoplasm of 3T3 cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/ml, fixed with 2% paraformaldehyde, washed with PBS, and incubated with ab2806 (1:80) for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

  • Anti-ADP Ribosylation Factor [1D9] antibody (ab2806) + Canine heart extract
  • Ab2806 immunoprecipitating ADP Ribosylation Factor from HeLa cells. 10ul of antibody were added to 500ul of HeLa cell lysate (at 1.5mg/ml) and incubated overnight before precipitation with Protein G. Negative control lysate received the same treatment except no antibody was added. ab2806 was used at 1/500 in the western blotting stage and the secondary antibody was an HRP conjugated anti-mouse IgG. The membrane was finally stained with amido black.

    Lane 1: negative control

    Lane 2: ab2806

    This image is courtesy of an Abreview submitted by Denis Krndija submitted on 5 January 2006.

    See Abreview

  • All lanes : Anti-ADP Ribosylation Factor [1D9] antibody (ab2806) at 1/500 dilution

    Lane 1 : Lysates prepared from human Huh-7 cells
    Lane 2 : Lysates prepared from human Huh-7 cells

    Lysates/proteins at 15000 cells per lane.

    Secondary
    HRP-conjugated sheep polyclonal to mouse IgG at 1/10000 dilution

    Performed under reducing conditions.

    Observed band size : 21 kDa (why is the actual band size different from the predicted?)


    Exposure time : 4 minutes

    This image is a courtesy of Anonymous Abreview

    See Abreview

  • Overlay histogram showing HeLA cells stained with ab2806 (red line). The cells were fixed with 100% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab2806, 1/20 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human colon carcinoma tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing ADP-Ribosylation Factor ab2806 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human tonsil tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:50 with a mouse monoclonal antibody recognizing ADP-Ribosylation Factor ab2806 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human liver tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:50 with a mouse monoclonal antibody recognizing ADP-Ribosylation Factor ab2806 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

References for Anti-ADP Ribosylation Factor [1D9] antibody (ab2806)

This product has been referenced in:
  • Quilty D  et al. Arf activation at the Golgi is modulated by feed-forward stimulation of the exchange factor GBF1. J Cell Sci 127:354-64 (2014). Read more (PubMed: 24213530) »
  • Kametaka S  et al. Functional characterization of protein-sorting machineries at the trans-Golgi network in Drosophila melanogaster. J Cell Sci 123:460-71 (2010). WB ; Fruit fly (Drosophila melanogaster) . Read more (PubMed: 20067992) »

See all 9 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Sample Fruit fly (Drosophila melanogaster) Cell (Whole embryo)
Specification Whole embryo
Fixative see additional notes
Permeabilization Yes - Heptane
Blocking step Normal Goat Serum (NGS) as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 20°C
Username

Mr. Donghoon Lee

Verified customer

Submitted Aug 02 2010

Application Western blot
Sample Human Cell lysate - whole cell (huh-7)
Loading amount 15000 cells
Specification huh-7
Gel Running Conditions Reduced Denaturing
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Jul 29 2009

Application Western blot
Sample Rat Cell lysate - other (NRK cell Postnuclear supernatant)
Loading amount 50 µg
Specification NRK cell Postnuclear supernatant
Gel Running Conditions Reduced Denaturing
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Sep 18 2007

Thank you for your enquiry. Unfortunately we have not done testing to quantify the relative amounts of ARF isotypes from human whole cell lysate at this time. I apologize for any inconvenience this may cause you. I do have one article that specifica...

Read More
Application Immunoprecipitation
Sample Human Cell lysate - whole cell (HeLa cells)
Specification HeLa cells
Immuno-precipitation step Protein G
Username

Mr. Denis Krndija

Verified customer

Submitted Jan 05 2006

Application Immunocytochemistry
Sample Human Cell (HeLa)
Specification HeLa
Fixative Paraformaldehyde
Blocking step Other as blocking agent for 30 minute(s) · Concentration: 0.2%
Username

Mr. Denis Krndija

Verified customer

Submitted Oct 26 2005

Thank you for your enquiry. After reviewing the data, the molecular weight markers on this data sheet appear to be incorrect. The band should be between the 14 KDa and 20 KDa, as indicated by Swiss Prot. There are many different forms of ARF and all...

Read More

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