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FANCD2 Recombinant Rabbit Monoclonal Antibody [SN73-08]

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Specification

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Catalog# ET1611-67

FANCD2 Recombinant Rabbit Monoclonal Antibody [SN73-08]

Application

  • WB

  • IF-Cell

  • IF-Tissue

  • IHC-P

Reactivity

  • Human

  • Mouse

  • Rat

Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

FANCD2 Recombinant Rabbit Monoclonal Antibody [SN73-08]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within Human FANCD2 aa 191-240 / 1,451.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IF-Cell, IF-Tissue, IHC-P

Molecular Weight

Predicted band size: 164 kDa

Positive Control

HeLa cell lysate, K-562 cell lysate, RAW264.7 cell lysate, C6 cell lysate, Mouse testis tissue lysate, Rat testis tissue lysate, HeLa, C6, mouse testis tissue, rat stomach tissue, rat testis tissue, mouse stomach tissue.

Conjugation

unconjugated

Clone Number

SN73-08

RRID

AB_3070046

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:500-1:2,000

  • IF-Cell

  • 1:50-1:100

  • IF-Tissue

  • 1:100-1:500

  • IHC-P

  • 1:50-1:200

Target

Function

Fanconi anemia (FA) is an autosomal recessive disorder character-ized by bone marrow failure, birth defects and chromosomal instability. At the cellular level, FA is characterized by spontaneous chromosomal breakage and a unique hypersensitivity to DNA cross-linking agents. At least 8 complementation groups (A-G) have been identified and 6 FA genes (for subtypes A, C, D2, E, F and G) have been cloned. The FA proteins lack sequence homologies or motifs that could point to a molecular function. Phosphorylation of FANC (Fanconi anemia complementation group) proteins are thought to be important for the function of the FA pathway. Several FA proteins, including FANCA, FANCC, FANCF, and FANCG, interact in a nuclear complex, and this complex is required for the activation (monoubiquitination) of the downstream FANCD2 protein. When monoubiquitinated, the FANCD2 protein co-localizes with the breast cancer susceptibility protein BRCA1 in DNA damage induced foci. In male meiosis, FANCD2 also co-localizes with BRCA1 at synaptonemal complexes. The human FANCD2 gene maps to chromosome 3p25.3, contains 44 exons and encodes a 1,451-amino acid nuclear protein that exists as 2 protein isoforms.

Background References

1. Jiang Q et al. MERIT40 cooperates with BRCA2 to resolve DNA interstrand cross-links. Genes Dev 29:1955-68 (2015).

2. Xu W et al. TXNL1-XRCC1 pathway regulates cisplatin-induced cell death and contributes to resistance in human gastric cancer. Cell Death Dis 5:e1055 (2014).

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.

Post-translational Modification

Monoubiquitinated on Lys-561 during S phase and upon genotoxic stress by FANCL in complex with E2 ligases UBE2T or UBE2W (isoform 1 and isoform 2). Deubiquitinated by USP1 as cells enter G2/M, or once DNA repair is completed. Monoubiquitination requires the joint intervention of the FANC core complex, including FANCA, FANCB, FANCC, FANCE, FANCF, FANCG, and FANCM, and proteins involved in cell cycle checkpoints and DNA repair, including RPA1, ATR, CHEK1 and BRCA1, 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.

Subcellular Location

Nucleus, Cytoplasm.

UNIPROT

SwissProt: Q9BXW9 Human

SwissProt: Q80V62 Mouse

SwissProt: Q6IV68 Rat

Synonyms

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

Expand

Images

  • Western blot analysis of FANCD2 on different lysates with Rabbit anti-FANCD2 antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/2,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: K-562 cell lysate<br />Lane 3: RAW264.7 cell lysate<br />Lane 4: C6 cell lysate<br />Lane 5: Mouse testis tissue lysate<br />Lane 6: Rat testis tissue lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 164 kDa<br />Observed band size: 164 kDa<br /><br />Exposure time: Lane 1: 8 seconds; Lane 2-3: 30 seconds; Lane 4-6: 3 minutes; ECL: K1802;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of FANCD2 on different lysates with Rabbit anti-FANCD2 antibody (ET1611-67) at 1/2,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: K-562 cell lysate
    Lane 3: RAW264.7 cell lysate
    Lane 4: C6 cell lysate
    Lane 5: Mouse testis tissue lysate
    Lane 6: Rat testis tissue lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 164 kDa
    Observed band size: 164 kDa

    Exposure time: Lane 1: 8 seconds; Lane 2-3: 30 seconds; Lane 4-6: 3 minutes; ECL: K1802;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1611-67) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunocytochemistry analysis of HeLa cells labeling FANCD2 with Rabbit anti-FANCD2 antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-FANCD2 antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/HA601187" style="font-weight: bold;text-decoration: underline;">HA601187</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of HeLa cells labeling FANCD2 with Rabbit anti-FANCD2 antibody (ET1611-67) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-FANCD2 antibody (ET1611-67) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Immunocytochemistry analysis of C6 cells labeling FANCD2 with Rabbit anti-FANCD2 antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/50 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-FANCD2 antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>) at 1/50 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/HA601187" style="font-weight: bold;text-decoration: underline;">HA601187</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of C6 cells labeling FANCD2 with Rabbit anti-FANCD2 antibody (ET1611-67) at 1/50 dilution.

    Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-FANCD2 antibody (ET1611-67) at 1/50 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Immunohistochemical analysis of paraffin-embedded mouse testis tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded mouse testis tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1611-67, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded rat stomach tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded rat stomach tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1611-67, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded rat testis tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded rat testis tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1611-67, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded mouse stomach tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1611-67" style="font-weight: bold;text-decoration: underline;">ET1611-67</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded mouse stomach tissue using anti-FANCD2 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1611-67, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

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