リコンビナント
RabMAb

Anti-NEDD8 抗体 [Y297] (ab81264)

製品の概要

  • 製品名
    Anti-NEDD8 antibody [Y297]
    NEDD8 一次抗体 製品一覧
  • 製品の詳細
    Rabbit monoclonal [Y297] to NEDD8
  • 特異性
    ab81264 detects NEDD8 bound to cullins proteins (neddylated cullins) as well as free NEDD8.
  • アプリケーション
    適用あり: Flow Cyt, WB, IHC-P, ICC/IF, IPmore details
  • 種交差性
    交差種: Mouse, Rat, Human
  • 免疫原

    corresponding to Human NEDD8 (N terminal).

  • ポジティブ・コントロール
    • Hela and K562 cell lysates and human lung carcinoma. FC: A-673
  • 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

    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.

製品の特性

  • 製品の状態
    Liquid
  • 保存方法
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Stable for 12 months at -20°C.
  • バッファー
    PBS 49%,Sodium azide 0.01%,Glycerol 50%,BSA 0.05%
  • Concentration information loading...
  • 精製度
    Protein A purified
  • ポリ/モノ
    モノクローナル
  • クローン名
    Y297
  • アイソタイプ
    IgG
  • 研究分野

アプリケーション

Our Abpromise guarantee covers the use of ab81264 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
Flow Cyt 1/300.
WB 1/1000 - 1/10000. Detects a band of approximately 9 kDa (predicted molecular weight: 9 kDa).
IHC-P 1/100 - 1/500.
ICC/IF 1/100 - 1/250.
IP 1/20 - 1/50.

ターゲット情報

  • 機能
    Ubiquitin-like protein which plays an important role in cell cycle control and embryogenesis. Covalent attachment to its substrates requires prior activation by the E1 complex UBE1C-APPBP1 and linkage to the E2 enzyme UBE2M. Attachment of NEDD8 to cullins activates their associated E3 ubiquitin ligase activity, and thus promotes polyubiquitination and proteasomal degradation of cyclins and other regulatory proteins.
  • 組織特異性
    Highly expressed in heart, skeletal muscle, spleen, thymus, prostate, testis, ovary, colon and leukocytes.
  • 配列類似性
    Belongs to the ubiquitin family.
  • 翻訳後修飾
    Cleavage of precursor form by UCHL3 or SENP8 is necessary for function.
  • 細胞内局在
    Nucleus. Mainly nuclear.
  • Information by UniProt
  • 参照データベース
  • 別名
    • FLJ43224 antibody
    • MGC104393 antibody
    • MGC125896 antibody
    • MGC125897 antibody
    • NED8 antibody
    • NEDD 8 antibody
    • NEDD-8 antibody
    • Nedd8 antibody
    • NEDD8_HUMAN antibody
    • Neddylin antibody
    • Neural precursor cell expressed developmentally down regulated 8 antibody
    • Neural precursor cell expressed developmentally down regulated gene 8 antibody
    • Neural precursor cell expressed developmentally down-regulated protein 8 antibody
    • Rub1 antibody
    • Ubiquitin like protein Nedd 8 antibody
    • Ubiquitin like protein Nedd8 antibody
    • Ubiquitin-like protein Nedd8 antibody
    see all

画像

  • All lanes : Anti-NEDD8 antibody [Y297] (ab81264) at 1/1000 dilution (Unpurified)

    Lane 1 : C6 Cell Lysate
    Lane 2 : Raw264.7 Cell Lysate
    Lane 3 : PC-12 Cell Lysate
    Lane 4 : 3T3 Cell Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

    Predicted band size : 9 kDa
  • Immunohistochemical staining of paraffin embedded human cervical carcinoma with purified ab81264 at a working dilution of 1/400. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
  • Immunofluorescence staining of HeLa cells with purified ab81264 at a working dilution of 1/100, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab81264 was used at a dilution of 1/500 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

  • Flow cytometry analysis of A-673 (Human muscle Ewing's Sarcoma) cells labeling NEDD8 (red) with purified ab81264 at a 1/300 dilution (1ug/mL). 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 (Black) (ab172730). Blue (unlabeled control) - Cell without incubation with primary antibody and secondary antibody (Blue).

  • Anti-NEDD8 antibody [Y297] (ab81264) at 1/1000 dilution (purified) + PC-12 whole cell lysate at 10 µg

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

    Predicted band size : 9 kDa
    Observed band size : 12,90 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

    The band at 12kDa is free NEDD8, and at 90kDa is Neddylated cullins.

  • Anti-NEDD8 antibody [Y297] (ab81264) at 1/1000 dilution (purified) + K562 whole cell lysate at 10 µg

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

    Predicted band size : 9 kDa
    Observed band size : 12,90 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

    The band at 12kDa is free NEDD8, and at 90kDa is Neddylated cullins.

  • Anti-NEDD8 antibody [Y297] (ab81264) at 1/1000 dilution (purified) + HEK293 cell lysate at 10 µg

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

    Predicted band size : 9 kDa
    Observed band size : 12,90 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

    The band at 12kDa is free NEDD8, and at 90kDa is Neddylated cullins.

  • ab81264 (purified) at 1/20 immunoprecipitating NEDD8 in 10 μg K562 cell lysate (Lanes 1 and 2, observed at 10 and 90 kDa). Lane 3 - Rabbit monoclonal IgG (ab172730). For western blotting, HRP Veriblot for IP (ab131366) was used as the secondary antibody (1/10 000). Blocking buffer and concentration: 5% NFDM/TBST Dilution buffer and concentration: 5% NFDM/TBST
  • All lanes : Anti-NEDD8 antibody [Y297] (ab81264) at 1/500 dilution (Unpurified)

    Lane 1 : HeLa cell lysate
    Lane 2 : K562 cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP labelled goat anti-rabbit at 1/2000 dilution

    Predicted band size : 9 kDa
    Observed band size : 9 kDa
    Other Bands: 90kDa Neddylated cullins
  • Western blot analysis of immunoprecipitation pellet from HeLa cell lysate using unpurified ab81264 at 1/500 dilution. Band at 90 kDa shows Neddylated cullins, 50 kDa the IgG fraaction and 9 kDa Free NEDD8.

  • Unpurified ab81264, at 1/100 dilution, staining NEDD8 in human lung carcinoma tissue by Immunohistochemistry using formalin-fixed, paraffin-embedded section.

参考文献

This product has been referenced in:
  • Lu R  et al. COPS5 amplification and overexpression confers tamoxifen-resistance in ERa-positive breast cancer by degradation of NCoR. Nat Commun 7:12044 (2016). Human . Read more (PubMed: 27375289) »
  • Song H  et al. MLN4924, a First-in-Class NEDD8-Activating Enzyme Inhibitor, Attenuates IFN-ß Production. J Immunol 196:3117-23 (2016). Read more (PubMed: 26895833) »

See all 5 Publications for this product

レビューと Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Zebrafish Tissue lysate - whole (whole embryo)
Gel Running Conditions
Non-reduced Denaturing (12%)
Loading amount
40 µg
Specification
whole embryo
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 Apr 06 2016

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

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