Overview

  • Product name

    Anti-FANCD2 antibody [EPR2302]
    See all FANCD2 primary antibodies
  • Description

    Rabbit monoclonal [EPR2302] to FANCD2
  • Host species

    Rabbit
  • Specificity

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

  • Tested applications

    Suitable for: Flow Cyt, IHC-P, ICC/IF, IP, WBmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human FANCD2 aa 200-300. The exact sequence is proprietary.

  • Positive control

    • WB: HAP1, HeLa and HEK293 cell lysates. IHC-P: Human tonsil tissue. ICC/IF: HAP1 and HeLa cells. Flow Cyt: Jurkat cells. IP: HeLa cell lysate
  • 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 long term. Avoid freeze / thaw cycle.
  • 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

    EPR2302
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab108928 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
Flow Cyt 1/200.
IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

 

 

See IHC antigen retrieval protocols.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

ICC/IF 1/250 - 1/500.
IP 1/10 - 1/100.
WB 1/1000 - 1/10000. Detects a band of approximately 155 kDa (predicted molecular weight: 166 kDa).

Target

  • Function

    Required for maintenance of chromosomal stability. Promotes accurate and efficient pairing of homologs during meiosis. Involved in the repair of DNA double-strand breaks, both by homologous recombination and single-strand annealing. May participate in S phase and G2 phase checkpoint activation upon DNA damage. Promotes BRCA2/FANCD1 loading onto damaged chromatin. May also be involved in B-cell immunoglobulin isotype switching.
  • Tissue specificity

    Highly expressed in germinal center cells of the spleen, tonsil, and reactive lymph nodes, and in the proliferating basal layer of squamous epithelium of tonsil, esophagus, oropharynx, larynx and cervix. Expressed in cytotrophoblastic cells of the placenta and exocrine cells of the pancreas (at protein level). Highly expressed in testis, where expression is restricted to maturing spermatocytes.
  • Involvement in disease

    Defects in FANCD2 are a cause of Fanconi anemia complementation group D type 2 (FANCD2) [MIM:227646]. It is a disorder affecting all bone marrow elements and resulting in anemia, leukopenia and thrombopenia. It is associated with cardiac, renal and limb malformations, dermal pigmentary changes, and a predisposition to the development of malignancies. At the cellular level it is associated with hypersensitivity to DNA-damaging agents, chromosomal instability (increased chromosome breakage) and defective DNA repair.
  • Developmental stage

    Highly expressed in fetal oocytes, and in hematopoietic cells of the fetal liver and bone marrow (at protein level).
  • Domain

    The C-terminal 24 residues of isoform 2 are required for its function.
  • Post-translational
    modifications

    Monoubiquitinated on Lys-561 during S phase and upon genotoxic stress (isoform 1 and isoform 2). Deubiquitinated by USP1 as cells enter G2/M, or once DNA repair is completed. Monoubiquitination requires the FANCA-FANCB-FANCC-FANCE-FANCF-FANCG-FANCM complex, RPA1 and ATR, and is mediated by FANCL/PHF9. Ubiquitination is required for binding to chromatin, interaction with BRCA1, BRCA2 and MTMR15/FAN1, DNA repair, and normal cell cycle progression, but not for phosphorylation on Ser-222 or interaction with MEN1.
    Phosphorylated in response to various genotoxic stresses by ATM and/or ATR. Upon ionizing radiation, phosphorylated by ATM on Ser-222 and Ser-1404. Phosphorylation on Ser-222 is required for S-phase checkpoint activation, but not for ubiquitination, foci formation, or DNA repair. In contrast, phosphorylation by ATR on other sites may be required for ubiquitination and foci formation.
  • Cellular localization

    Nucleus. Concentrates in nuclear foci during S phase and upon genotoxic stress.
  • Information by UniProt
  • Database links

  • Alternative names

    • DKFZp762A223 antibody
    • FA 4 antibody
    • FA D2 antibody
    • FA4 antibody
    • FAC D2 antibody
    • FACD 2 antibody
    • FACD antibody
    • FACD2 antibody
    • FACD2_HUMAN antibody
    • FAD antibody
    • FAD2 antibody
    • FANC D2 antibody
    • FANCD 2 antibody
    • FANCD antibody
    • FANCD2 antibody
    • FANCONI ANEMIA COMPLEMENTATION GROUP D antibody
    • Fanconi anemia complementation group D2 antibody
    • Fanconi anemia group D2 protein antibody
    • FANCONI PANCYTOPENIA TYPE 4 antibody
    • FLJ23826 antibody
    • OTTHUMP00000158853 antibody
    • OTTHUMP00000207925 antibody
    • Protein FACD2 antibody
    • Type 4 Fanconi pancytopenia antibody
    see all

Images

  • All lanes : Anti-FANCD2 antibody [EPR2302] (ab108928) at 1/1000 dilution (Purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates
    Lane 3 : C6 (Rat glial tumor glial cell) whole cell lysates
    Lane 4 : RAW264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
    Lane 5 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size: 166 kDa
    Observed band size: 165 kDa
    why is the actual band size different from the predicted?

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: FANCD2 knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (20 µg)
    Lane 4: HEK293 cell lysate (20 µg)
    Lanes 1 - 4: Merged signal (red and green). Green - ab108928 (unpurified) observed at 165 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab108928 was shown to specifically react with FANCD2 when FANCD2 knockout samples were used. Wild-type and FANCD2 knockout samples were subjected to SDS-PAGE. ab108928 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 1/10 000 dilution for 1 h at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human tonsil tissue sections labeling FANCD2 with purified ab108928 at 1/50 dilution (3.6 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
  • Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling FANCD2 with purified ab108928 at 1/500 dilution (0.4 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. 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.

  • ab108928 (purified) at 1/20 dilution (1ug) immunoprecipitating FANCD2 in HeLa whole cell lysates.
    Lane 1: HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates 10ug
    Lane 2 (+): ab108928 & HeLa whole cell lysates
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab108928 in HeLa whole cell lysates
    For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

  • Flow cytometry analysis of Jurkat (human acute T cell leukemia) cells labeling FANCD2 (red) with ab108928 (purified) at a 1/200 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with primary and secondary antibodies.

  • ab108928 (unpurified) staining FANCD2 in wild-type HAP1 cells (top panel) and FANCD2 knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10 minutes), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1 hour. The cells were then incubated with ab108928 at 1/250 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1 hour with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

  • Immunohistochemical staining of paraffin-embedded human tonsil tissue using ab108928 (unpurified) at a dilution of 1/100.

    Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

  • ab108928 (unpurified) staining FANCD2 in human U2OS osteosarcoma cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton X-100 and blocked with 10% goat serum for 1 hour at 20°C. Samples were incubated with primary antibody (1/300) for 18 hours at 4°C. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG monoclonal (1/1000) was used as the secondary antibody.

    See Abreview

  • HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained for FANCD2 (green) using ab108928 (unpurified) (1/250 dilution) in ICC/IF.

References

This product has been referenced in:

  • Spriggs CC  et al. Expression of HPV-induced DNA Damage Repair Factors Correlates With CIN Progression. Int J Gynecol Pathol 38:1-10 (2019). Read more (PubMed: 29995652) »
  • Yan W  et al. Inhibition of SKP2 Activity Impaired ATM-Mediated DNA Repair and Enhanced Sensitivity of Cisplatin-Resistant Mantle Cell Lymphoma Cells. Cancer Biother Radiopharm N/A:N/A (2019). Read more (PubMed: 31025879) »
See all 20 Publications for this product

Customer reviews and Q&As

1-4 of 4 Abreviews or Q&A

Application
Western blot
Sample
Human Cell lysate - whole cell (293T)
Gel Running Conditions
Non-reduced Non-Denaturing (Native)
Loading amount
60 µg
Treatment
MMC 2H
Specification
293T
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C

Abcam user community

Verified customer

Submitted Jul 31 2018

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C
Sample
Human Cell (u2os osteosarcoma)
Specification
u2os osteosarcoma
Permeabilization
Yes - 0.5% triton
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Apr 25 2014

Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing (6%)
Sample
Mouse Cell lysate - whole cell (Mouse Pancreatic cancer)
Specification
Mouse Pancreatic cancer
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 18 2014

Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (6%)
Sample
Human Tissue lysate - whole (U2OS osteosarcoma)
Specification
U2OS osteosarcoma
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Mar 04 2014

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