製品の概要

  • 製品名

    Anti-Rad21 antibody [EPR22506-15]
    Rad21 一次抗体 製品一覧
  • 製品の詳細

    Rabbit monoclonal [EPR22506-15] to Rad21
  • 由来種

    Rabbit
  • アプリケーション

    適用あり: WB, IHC-P, ICC/IF, Flow Cyt, ChIP, IPmore details
  • 種交差性

    交差種: Mouse, Rat, Human
  • 免疫原

    Synthetic peptide within Human Rad21 aa 300-400. The exact sequence is proprietary.
    Database link: O60216

  • ポジティブ・コントロール

    • WB: Human heart and fetal kidney tissue lysate. Rat brain and mouse brain and heart tissue lysate. HeLa, MCF7, Raw264.7, NIH/3T3 and C6 cell lysate. IHC-P: Human breast carcinoma, stomach, mouse stomach and rat colon tissue. ICC/IF: HeLa and NIH/3T3 cells. Flow: HeLa and NIH/3T3 cells. IP: HeLa whole cell lysate.
  • 特記事項

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

    This product is a recombinant rabbit monoclonal antibody.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab217678 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
WB 1/1000. Predicted molecular weight: 72 kDa.
IHC-P 1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

ICC/IF 1/100.
Flow Cyt 1/500.
ChIP Use at an assay dependent concentration.

5 µg

IP 1/30.

ターゲット情報

  • 機能

    Cleavable component of the cohesin complex, involved in chromosome cohesion during cell cycle, in DNA repair, and in apoptosis. The cohesin complex is required for the cohesion of sister chromatids after DNA replication. The cohesin complex apparently forms a large proteinaceous ring within which sister chromatids can be trapped. At metaphase-anaphase transition, this protein is cleaved by separase/ESPL1 and dissociates from chromatin, allowing sister chromatids to segregate. The cohesin complex may also play a role in spindle pole assembly during mitosis. Also plays a role in apoptosis, via its cleavage by caspase-3/CASP3 or caspase-7/CASP7 during early steps of apoptosis: the C-terminal 64 kDa cleavage product may act as a nuclear signal to initiate cytoplasmic events involved in the apoptotic pathway.
  • 配列類似性

    Belongs to the rad21 family.
  • ドメイン

    The C-terminal part associates with the head of SMC1A, while the N-terminal part binds to the head of SMC3.
  • 翻訳後修飾

    Cleaved by separase/ESPL1 at the onset of anaphase. Cleaved by caspase-3 and caspase-7 at the beginning of apoptosis. The cleavage by ESPL1 and caspase-3 take place at different sites.
    Phosphorylated; becomes hyperphosphorylated in M phase of cell cycle. The large dissociation of cohesin from chromosome arms during prophase may be partly due to its phosphorylation by PLK.
  • 細胞内局在

    Nucleus. Chromosome. Chromosome > centromere. Associates with chromatin. Before prophase it is scattered along chromosome arms. During prophase, most of cohesin complexes dissociate from chromatin probably because of phosphorylation by PLK, except at centromeres, where cohesin complexes remain. At anaphase, it is cleaved by separase/ESPL1, leading to the dissociation of the complex from chromosomes, allowing chromosome separation. Once cleaved by caspase-3, the C-terminal 64 kDa cleavage product translocates to the cytoplasm, where it may trigger apoptosis.
  • Information by UniProt
  • 参照データベース

  • 別名

    • CDLS4 antibody
    • Double-strand-break repair protein rad21 homolog antibody
    • hHR21 antibody
    • HR21 antibody
    • HRAD21 antibody
    • KIAA0078 antibody
    • MCD1 antibody
    • Nuclear matrix protein 1 antibody
    • NXP-1 antibody
    • NXP1 antibody
    • Protein involved in DNA double-strand break repair antibody
    • RAD21 antibody
    • RAD21 homolog (S. pombe) antibody
    • RAD21 homolog antibody
    • RAD21_HUMAN antibody
    • Scc1 antibody
    • SCC1 homolog antibody
    see all

画像

  • Chromatin was prepared from MCF7 cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min.

    The ChIP was performed with 25 µg of chromatin, 5 µg of ab217678 (red), and 20 µl of Protein A/G sepharose beads. 5 μg of rabbit normal IgG was added to the beads control (gray). The immunoprecipitated DNA was quantified by real time PCR (sybr green approach).

    Primers and probes are located in the first kb of the transcribed region.

    The observed enrichment profile of Rad21 binding is consistent with what has been described in the literature (PMID: 23145106)

     

  • All lanes : Anti-Rad21 antibody [EPR22506-15] (ab217678) at 1/1000 dilution

    Lane 1 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage), whole cell lysate
    Lane 2 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate
    Lane 3 : C6 (rat glial tumor glial cell), whole cell lysate whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 72 kDa
    Observed band size: 130-75 kDa
    why is the actual band size different from the predicted?



    Full length Rad21 is approximately 130kDa, cleaved Rad21 is approximately 75 kDa. The molecular weight observed are consistent with what has been described in the literature (PMID: 12417729, PMID 21876002).

    Exposure time: 7.7 seconds

    Blocking/diluting buffer and concentration: 5% NFDM/TBST

  • All lanes : Anti-Rad21 antibody [EPR22506-15] (ab217678) at 1/1000 dilution

    Lane 1 : Rat Brain tissue lysate
    Lane 2 : Mouse Brain tissue lysate
    Lane 3 : Mouse heart tissue lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 72 kDa
    Observed band size: 130-75 kDa why is the actual band size different from the predicted?



    Full length Rad21 is approximately 130kDa, cleaved Rad21 is approximately 75 kDa. The molecular weight observed are consistent with what has been described in the literature (PMID: 12417729, PMID 21876002).

    Blocking/Diluting buffer and concentration: 5% NFDM/TBST

    Exposure time: 3 minutes.

     

  • All lanes : Anti-Rad21 antibody [EPR22506-15] (ab217678) at 1/1000 dilution

    Lane 1 : Human heart tissue lysate
    Lane 2 : Human fetal kidney tissue lysate
    Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate
    Lane 4 : MCF7 (human breast adenocarcinoma epithelial cell), whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Lanes 1-2 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution
    Lanes 3-4 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 72 kDa
    Observed band size: 130-75 kDa why is the actual band size different from the predicted?



    Full length Rad21 is approximately 130 kDa, cleaved Rad21 is approximately 75 kDa. The molecular weight observed are consistent with what has been described in the literature (PMID: 12417729, PMID 21876002).

    Blocking/Diluting buffer and concentration: 5% NFDM/TBST

    Exposure times:

    Lanes1-2: 37 seconds

    Lane 3: 5.5 seconds

    Lane 4: 48 seconds

  • Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling Rad21 using ab217678 at 1/1000 dilution for 30 minutes at room temperature, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on rat colon (PMID: 25575569) is observed. Counterstained with Hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is  a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemical analysis of paraffin-embedded mouse stomach tissue labeling Rad21 using ab217678 at 1/1000 dilution for 30 minutes at room temperature, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on mouse stomach (PMID: 25575569) is observed. Counterstained with Hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is  a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemical analysis of paraffin-embedded human stomach tissue labeling Rad21 using ab217678 at 1/1000 dilution for 30 minutes at room temperature, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on human stomach (PMID: 25575569) is observed. Counterstained with Hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is  a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

     

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue labeling Rad21 using ab217678 at 1/1000 dilution for 30 minutes at room temperature, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on the human breast carcinoma (PMID: 21255398) is observed. Counterstained with Hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is  a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

     

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Rad21 was immunoprecipitated from 0.35mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with ab217678 at 1/30 dilution. Western blot was performed on the immunoprecipitate using ab217678 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used as the secondary antibody at 1/5000 dilution.

    Lane 1: HeLa whole cell lysate 10µg (input)
    Lane 2: ab217678 IP in HeLa whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab217678 in HeLa whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 3 seconds.

    Faint band below 75kDa in lane 2 could be Rad21 cleavage product as described in the literature (PMID: 28854249).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed NIH/3T3 (mouse embryonic fibroblast) cell line labeling Rad21 using ab217678 at 1/500 dilution (red) compared with a Rabbit monoclonal IgG (ab172730, Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (human cervix adenocarcinoma epithelial cell) cell line labeling Rad21 using ab217678 at 1/500 dilution (red) compared with a Rabbit monoclonal IgG (ab172730, Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labeling Rad21 with ab217678 at 1/100 dilution, followed by an AlexaFluor®488 Goat anti-Rabbit secondary antibody (ab150077) at 1/1000 dilution (green). Confocal image showing nuclear staining in NIH/3T3 cell line is observed. The nuvlear counterstain is DAPI (Blue). Tubulin is counterstained using Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is an AlexaFluor®488 Goat anti-Rabbit secondary (ab150077) at 1/1000 dilution.

     

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labeling Rad21 with ab217678 at 1/100 dilution, followed by an AlexaFluor®488 Goat anti-Rabbit secondary antibody (ab150077) at 1/1000 dilution (green). Confocal image showing nuclear staining in HeLa cell line is observed. The nuvlear counterstain is DAPI (Blue). Tubulin is counterstained using Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is an AlexaFluor®488 Goat anti-Rabbit secondary (ab150077) at 1/1000 dilution.

     

参考文献

ab217678 has not yet been referenced specifically in any publications.

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