製品の概要

  • 製品名
    Anti-ABCA1 antibody [HJ1]
    ABCA1 一次抗体 製品一覧
  • 製品の詳細
    Mouse monoclonal [HJ1] to ABCA1
  • アプリケーション
    適用あり: WB, Flow Cyt, IHC-Pmore details
  • 種交差性
    交差種: Mouse, Rat, Human
  • 免疫原

    Recombinant fragment corresponding to ABCA1 (N terminal). 50 kDa N-terminal extracellular loop of ABCA1
    Database link: O95477

  • ポジティブ・コントロール
    • In Western Blot, ab66217 gave a positive signal in the following lysates: Rat Brain Tissue; Mouse Brain Tissue; Rat Liver Tissue; Mouse Liver Tissue; HepG2 Whole Cell. In IHC, this antibody gave a positive result in Human Liver FFPE Tissue.
  • 特記事項

    This monoclonal antibody to ABCA1 has been knockout validated in Western blot. The expected band for ABCA1 was observed in wild type cells and the band was not seen in knockout cells, although other non-specific bands were also seen. The data are shown on this datasheet.

    This antibody clone is manufactured by Abcam.

    If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

アプリケーション

Our Abpromise guarantee covers the use of ab66217 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 Use a concentration of 1 µg/ml. Detects a band of approximately 254 kDa (predicted molecular weight: 254 kDa).

Abcam recommends using 3% Milk as the blocking agent.

Flow Cyt Use 2µg for 106 cells.

(methanol fixed cells)

ab170192 - Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.

IHC-P Use a concentration of 0.2 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

ターゲット情報

  • 機能
    cAMP-dependent and sulfonylurea-sensitive anion transporter. Key gatekeeper influencing intracellular cholesterol transport.
  • 組織特異性
    Widely expressed, but most abundant in macrophages.
  • 関連疾患
    Defects in ABCA1 are a cause of high density lipoprotein deficiency type 1 (HDLD1) [MIM:205400]; also known as analphalipoproteinemia or Tangier disease (TGD). HDLD1 is a recessive disorder characterized by absence of high density lipoprotein (HDL) cholesterol from plasma, accumulation of cholesteryl esters, premature coronary artery disease (CAD), hepatosplenomegaly, recurrent peripheral neuropathy and progressive muscle wasting and weakness.
    Defects in ABCA1 are a cause of high density lipoprotein deficiency type 2 (HDLD2) [MIM:604091]; also known as familial hypoalphalipoproteinemia (FHA). HDLD2 is inherited as autosomal dominant trait. It is characterized by moderately low HDL cholesterol, predilection toward premature coronary artery disease (CAD) and a reduction in cellular cholesterol efflux.
  • 配列類似性
    Belongs to the ABC transporter superfamily. ABCA family.
    Contains 2 ABC transporter domains.
  • ドメイン
    Multifunctional polypeptide with two homologous halves, each containing an hydrophobic membrane-anchoring domain and an ATP binding cassette (ABC) domain.
  • 翻訳後修飾
    Phosphorylation on Ser-2054 regulates phospholipid efflux.
    Palmitoylation by DHHC8 is essential for membrane localization.
  • 細胞内局在
    Membrane.
  • Information by UniProt
  • 参照データベース
  • 別名
    • ABC 1 antibody
    • ABC Transporter 1 antibody
    • ABC-1 antibody
    • ABC1 antibody
    • ABCA 1 antibody
    • ABCA1 antibody
    • ABCA1_HUMAN antibody
    • ATP binding Cassette 1 antibody
    • ATP binding cassette sub family A ABC1 member 1 antibody
    • ATP binding cassette sub family A member 1 antibody
    • ATP binding Cassette Transporter 1 antibody
    • ATP-binding cassette 1 antibody
    • ATP-binding cassette sub-family A member 1 antibody
    • ATP-binding cassette transporter 1 antibody
    • CERP antibody
    • Cholesterol efflux regulatory protein antibody
    • FLJ14958 antibody
    • HDLDT1 antibody
    • Membrane bound antibody
    • MGC164864 antibody
    • MGC165011 antibody
    • TD antibody
    • TGD antibody
    see all

画像

  • ab66217 (4µg/ml) staining ABCA1 in human liver using an automated system (DAKO Autostainer Plus). Using this protocol there is moderate cell membrane staining throughout the liver parenchyma.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
  • IHC image of ABCA1 staining in human liver formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab66217, 0.2µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
  • Overlay histogram showing HepG2 cells stained with ab66217 (red line). The cells were fixed with methanol (5 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab66217, 2µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in HepG2 cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.

    Please note that Abcam does not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.
  • All lanes : Anti-ABCA1 antibody [HJ1] (ab66217) at 1 µg/ml

    Lane 1 : Brain (Mouse) Tissue Lysate
    Lane 2 : Brain (Rat) Tissue Lysate
    Lane 3 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
    Lane 4 : Liver (Mouse) Tissue Lysate
    Lane 5 : Liver (Rat) Tissue Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat polyclonal Secondary Antibody to Mouse IgG - H&L (HRP), pre-adsorbed at 1/5000 dilution

    Performed under reducing conditions.

    Predicted band size : 254 kDa
    Observed band size : 254 kDa
    Additional bands at : 70 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 4 minutesAbcam recommends using milk as the blocking agent. This blot was produced using a 3-8% Tris Acetate gel under the TA buffer system. The gel was run at 150V for 60 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% Milk before being incubated with ab66217 overnight at 4°C. Antibody binding was detected using an anti-mouse antibody conjugated to HRP, and visualised using ECL development solution.


  • Predicted band size : 254 kDa

    Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: Empty
    Lane 3: ABCA1 knockout (KO) HAP1 whole cell lysate (20 µg)

     

    Lanes 1 - 3: Merged signal (red and green). Green - ab66217 observed at 240 kDa. Red - loading control, ab176560, observed at 50 kDa.

     

    ab66217 detected the expected band for ABCA1 in wild type HAP1 cells and the band was not seen in ABCA1 knockout HAP1 cells, although additional non-specific bands were also seen. Wild-type and ABCA1 knockout samples were subjected to SDS-PAGE. Ab66217 and ab176560 (Rabbit anti alpha Tubulin loading control) were incubated overnight at 4°C at 1 µg/ml and 1/10000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

参考文献

This product has been referenced in:

See all 4 Publications for this product

レビューと Q&A


Yes, I can send you as a one time exception a positive control lysate (rat liver (ab29413), http://www.abcam.com/liver-rat-whole-cell-lysate-normal-tissue-ab29413.html). This has been tested with ab66217 and gave strong bands.

Also I woul...

Read More
Application
Immunocytochemistry
Sample
Mouse Cultured Cells (RAW 264.7)
Specification
RAW 264.7
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton-X100 in 2% BSA for 15min
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 22°C
Username

Dr. Mahesh Shivananjappa

Verified customer

投稿 May 19 2012

Thank you for submitting your recent review of ab66217. We are sorry to see that the results were unsatisfactory. I'd like to follow up with you to offer a couple of suggestions that may help improve these results. However we do fully guarantee this an...

Read More
Application
Western blot
Sample
Human Cell lysate - other (Mononuclear Cell Preparations)
Loading amount
25 µg
Specification
Mononuclear Cell Preparations
Gel Running Conditions
Reduced Denaturing (4-15%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Dr. Mahesh Shivananjappa

Verified customer

投稿 Nov 23 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Flow Cytometry
Sample
Human Cell (Platelets)
Specification
Platelets
Preparation
Cell harvesting/tissue preparation method: PL were isolated spinning Platelet rich plasma on Histopaque
Sample buffer: PBS
Fixation
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton-X100 in 2% BSA for 15min
Gating Strategy
Platelets
Username

Abcam user community

Verified customer

投稿 Dec 22 2010

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

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