Overview

  • Product name
    Anti-RPA70 antibody [EPR3472]
    See all RPA70 primary antibodies
  • Description
    Rabbit monoclonal [EPR3472] to RPA70
  • Tested applications
    Suitable for: ICC/IF, WB, IP, IHC-P, Flow Cyt, RIPmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human RPA70 aa 1-100.

  • Positive control
    • WB: A549, HEK293 and HeLa cell lysates IHC-P: Human cervical squamous cell carcinoma tissue. ICC/IF: A549 cells. Flow Cyt: HeLa cells. IP: HeLa whole cell lysate.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

    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.

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

     

    Alternative versions available:

    Anti-RPA70 antibody (Alexa Fluor® 488) [EPR3472] (ab199097)
    Anti-RPA70 antibody (Alexa Fluor® 647) [EPR3472] (ab199240)

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

Properties

Applications

Our Abpromise guarantee covers the use of ab79398 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 1/100 - 1/250.
WB 1/1000. Detects a band of approximately 70 kDa (predicted molecular weight: 70 kDa).

For unpurified use at 1/2000 - 1/5000.

IP 1/10 - 1/20.
IHC-P 1/100 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See protocols (link: http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol).

Flow Cyt 1/80.

For unpurified use at 1/50.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

RIP Use at an assay dependent concentration. PubMed: 21957289

Target

  • Function
    Plays an essential role in several cellular processes in DNA metabolism including replication, recombination and DNA repair. Binds and subsequently stabilizes single-stranded DNA intermediates and thus prevents complementary DNA from reannealing.
    Functions as component of the alternative replication protein A complex (aRPA). aRPA binds single-stranded DNA and probably plays a role in DNA repair; it does not support chromosomal DNA replication and cell cycle progression through S-phase. In vitro, aRPA cannot promote efficient priming by DNA polymerase alpha but supports DNA polymerase delta synthesis in the presence of PCNA and replication factor C (RFC), the dual incision/excision reaction of nucleotide excision repair and RAD51-dependent strand exchange.
  • Sequence similarities
    Belongs to the replication factor A protein 1 family.
  • Post-translational
    modifications
    Phosphorylated upon DNA damage, probably by ATM or ATR.
    Sumoylated on lysine residues Lys-449 and Lys-577, with Lys-449 being the major site. Sumoylation promotes recruitment of RAD51 to the DNA damage foci to initiate DNA repair through homologous recombinaison. Desumoylated by SENP6.
  • Cellular localization
    Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • Dmrpa1 antibody
    • Drosophila Replication Protein A antibody
    • DRPA antibody
    • HSSB antibody
    • Human single stranded DNA binding protein antibody
    • MST075 antibody
    • MSTP075 antibody
    • p70 antibody
    • REPA1 antibody
    • Replication factor A antibody
    • Replication factor A protein 1 antibody
    • Replication protein A 70 kDa DNA-binding subunit antibody
    • Replication protein A 70kDa DNA binding subunit antibody
    • Replication protein A1 70kDa antibody
    • Replication protein A1 antibody
    • RF A antibody
    • RF-A protein 1 antibody
    • RFA antibody
    • RFA1_HUMAN antibody
    • RP A antibody
    • RP-A p70 antibody
    • RPA 70 antibody
    • RPA antibody
    • rpa1 antibody
    • Single stranded binding protein 70 antibody
    • Single-stranded DNA-binding protein antibody
    see all

Anti-RPA70 antibody [EPR3472] images

  • Anti-RPA70 antibody [EPR3472] (ab79398) at 1/1000 dilution (purified) + A549 cell lysate at 20 µg

    Secondary
    Peroxidase-conjugated goat anti-rabbit IgG, (H+L) at 1/1000 dilution

    Predicted band size : 70 kDa
    Observed band size : 70 kDa

    Blocking and dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-RPA70 antibody [EPR3472] (ab79398) at 1/4000 dilution (purified)

    Lane 1 : HEK293 cell lysate
    Lane 2 : HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Peroxidase-conjugated goat anti-rabbit IgG, (H+L) at 1/1000 dilution

    Predicted band size : 70 kDa
    Observed band size : 70 kDa

    Blocking and dilution buffer: 5% NFDM/TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervix carcinoma tissue labelling RPA70 with purified ab79398 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunocytochemistry/Immunofluorescence analysis of A549 cells labelling RPA70 with purified ab79398 at 1/200. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500) were also used.

    Control 1: primary antibody (1/200) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).

    Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).

  • Flow Cytometry analysis of HeLa cells labelling RPA70 with purified ab79398 at 1/80 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

  • ab79398 (purified) at 1/20 immunoprecipitating RPA70 in HeLa whole cell lysate. 

    Lane 1 (input): HeLa whole cell lysate (10µg)

    Lane 2 (+): ab79398 + HeLa whole cell lysate (10µg).

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab79398 in HeLa whole cell lysate.

    For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-RPA70 antibody [EPR3472] (ab79398) at 1/5000 dilution (unpurified)

    Lane 1 : A549 cell lysate
    Lane 2 : HeLa cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP-conjugated goat anti-rabbit IgG at 1/2000 dilution

    Predicted band size : 70 kDa
    Observed band size : 70 kDa
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical squamous cell carcinoma labelling RPA70 with unpurified ab79398 at a dilution of 1/100.

  • Unpurified ab79398 staining RPA70 in U2OS cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton X-100 in PBS and blocked with 2% BSA for 1 hour at 25°C. Samples were incubated with primary antibody (1/500 in PBS + 0.5% Tween-20) for 2 hours at 25°C. A Cy3®-conjugated goat anti-rabbit IgG monoclonal (1/250) was used as the secondary antibody.

    See Abreview

  • Overlay histogram showing HeLa cells stained with unpurified ab79398 (red line). The cells were fixed with 80% 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 (unpurified ab79398, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

References for Anti-RPA70 antibody [EPR3472] (ab79398)

This product has been referenced in:
  • Bélanger F  et al. Mutations in Replicative Stress Response Pathways Are Associated with S Phase-specific Defects in Nucleotide Excision Repair. J Biol Chem 291:522-37 (2016). WB ; Human . Read more (PubMed: 26578521) »
  • Guven M  et al. Photosensitized UVA-Induced Cross-Linking between Human DNA Repair and Replication Proteins and DNA Revealed by Proteomic Analysis. J Proteome Res 15:4612-4623 (2016). WB ; Human . Read more (PubMed: 27654267) »

See all 8 Publications for this product

Product Wall

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
Sample
Human Cell (U2OS)
Specification
U2OS
Permeabilization
Yes - 0.5% Triton /PBS
Fixative
Paraformaldehyde
Username

Dr. Remi Buisson

Verified customer

Submitted Nov 21 2014

Application
Immunoprecipitation
Immuno-precipitation step
Protein G
Sample
Human Cell lysate - nuclear (Hela cells)
Specification
Hela cells
Total protein in input
200 µg
Username

Dr. Remi Buisson

Verified customer

Submitted Nov 20 2014

Application
Flow Cytometry
Fixation
Paraformaldehyde
Permeabilization
Yes - 1xPBS + 0.2% Triton X-100
Sample
Human Cell (Human osteosarcoma U2OS cells)
Specification
Human osteosarcoma U2OS cells
Gating Strategy
Single cells discriminated by DAPI staining
Preparation
Cell harvesting/tissue preparation method: Trypsinization
Sample buffer: 1xPBS + 1 mg/ml BSA + 1 microg/ml DAPI + 250 microg/ml RNase
Username

Abcam user community

Verified customer

Submitted Aug 26 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - nuclear (Hela Cells)
Loading amount
50 µg
Specification
Hela Cells
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Oct 03 2012

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

Sign up