Anti-AIF 抗体 - Mitochondrial Marker (ab1998)

Reviews (2)Q&A (5)References (31)

製品の概要

  • 製品名

    Anti-AIF antibody - Mitochondrial Marker
    AIF 一次抗体 製品一覧

  • 製品の詳細

    Rabbit polyclonal to AIF - Mitochondrial Marker

  • 由来種

    Rabbit

  • アプリケーション

    適用あり: WB, IHC-P, ICC/IFmore details

  • 種交差性

    交差種: Mouse, Rat, Human

  • 免疫原

    Synthetic peptide corresponding to Human AIF aa 517-531.
    Database link: O95831
    (Peptide available as ab7870)

  • ポジティブ･コントロール

    • K562 cell lysate

  • 特記事項

    Apoptosis Inducing Factor

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab1998 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 0.25 - 1 µg/ml. Detects a band of approximately 67 kDa (predicted molecular weight: 67 kDa). Can be blocked with AIF (internal) peptide (human).
IHC-P Use at an assay dependent concentration.
ICC/IF Use a concentration of 1 µg/ml.

ターゲット情報

  • 機能

    Probable oxidoreductase that has a dual role in controlling cellular life and death; during apoptosis, it is translocated from the mitochondria to the nucleus to function as a proapoptotic factor in a caspase-independent pathway, while in normal mitochondria, it functions as an antiapoptotic factor via its oxidoreductase activity. The soluble form (AIFsol) found in the nucleus induces 'parthanatos' i.e., caspase-independent fragmentation of chromosomal DNA. Interacts with EIF3G,and thereby inhibits the EIF3 machinery and protein synthesis, and activates casapse-7 to amplify apoptosis. Plays a critical role in caspase-independent, pyknotic cell death in hydrogen peroxide-exposed cells. Binds to DNA in a sequence-independent manner.

  • 関連疾患

    Defects in AIFM1 are the cause of combined oxidative phosphorylation deficiency type 6 (COXPD6) [MIM:300816]. It is a mitochondrial disease resulting in a neurodegenerative disorder characterized by psychomotor delay, hypotonia, areflexia, muscle weakness and wasting.

  • 配列類似性

    Belongs to the FAD-dependent oxidoreductase family.

  • 翻訳後修飾

    Under normal conditions, a 54-residue N-terminal segment is first proteolytically removed during or just after translocation into the mitochondrial intermembrane space (IMS) by the mitochondrial processing peptidase (MPP) to form the inner-membrane-anchored mature form (AIFmit). During apoptosis, it is further proteolytically processed at amino-acid position 101 leading to the generation of the mature form, which is confined to the mitochondrial IMS in a soluble form (AIFsol). AIFsol is released to the cytoplasm in response to specific death signals, and translocated to the nucleus, where it induces nuclear apoptosis in a caspase-independent manner.

  • 細胞内局在

    Mitochondrion intermembrane space. Mitochondrion inner membrane. Cytoplasm. Nucleus. Cytoplasm > perinuclear region. Proteolytic cleavage during or just after translocation into the mitochondrial intermembrane space (IMS) results in the formation of an inner-membrane-anchored mature form (AIFmit). During apoptosis, further proteolytic processing leads to a mature form, which is confined to the mitochondrial IMS in a soluble form (AIFsol). AIFsol is released to the cytoplasm in response to specific death signals, and translocated to the nucleus, where it induces nuclear apoptosis. Colocalizes with EIF3G in the nucleus and perinuclear region.
  • Information by UniProt

  • 参照データベース

    see all

  • 別名

    • AIFM1 antibody
    • AIFM1_HUMAN antibody
    • Apoptosis inducing factor 1, mitochondrial antibody
    • Apoptosis inducing factor antibody
    • Apoptosis inducing factor, mitochondrion associated, 1 antibody
    • Apoptosis-inducing factor 1 antibody
    • CMTX4 antibody
    • COWCK antibody
    • COXPD6 antibody
    • Harlequin antibody
    • Hq antibody
    • mAIF antibody
    • MGC111425 antibody
    • MGC5706 antibody
    • mitochondrial antibody
    • Neuropathy, axonal motor-sensory, with deafness and mental retardation antibody
    • neuropathy, axonal, motor-sensory with deafness and mental retardation (Cowchock syndrome) antibody
    • PDCD 8 antibody
    • PDCD8 antibody
    • Programmed cell death 8 (apoptosis inducing factor) antibody
    • Programmed cell death 8 antibody
    • Programmed cell death 8 isoform 1 antibody
    • Programmed cell death 8 isoform 2 antibody
    • Programmed cell death 8 isoform 3 antibody
    • Programmed cell death protein 8 antibody
    • Programmed cell death protein 8 mitochondrial antibody
    • Programmed cell death protein 8 mitochondrial precursor antibody
    • Programmed cell death protein 8 mitochondrial precursor antibody
    • Striatal apoptosis inducing factor antibody
    see all

画像

  • Western blot - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    Western blot - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    All lanes : Anti-AIF antibody - Mitochondrial Marker (ab1998) at 1 µg/ml

    Lane 1 : K562 cell lysate
    Lane 2 : Rat heart tissue lysate
    Lane 3 : Mouse heart tissue lysate

    Predicted band size: 67 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    Immunohistochemistry of AIF in human retina with anti-AIF (IN) at 10 µg/ml.
  • Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody - Mitochondrial Marker (ab1998)Image from Varecha M et al, J Biomed Sci. 2009 Jul 6;16:59, fig 2.
    ab1998 staining AIF in human U2OS cells by Immunocytochemistry/ Immunofluorescence.
    Samples were fixed using 4% paraformaldehyde.

    Left image shows fixed cells labeled with ab1998 (red) and cyclophilin A (green).
    Right image shows U2OS cell 6 hours after induction of apoptosis by 200 nM staurosporine.
    Note translocated of AIF to the nucleus upon induction of apoptosis.
  • Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody - Mitochondrial Marker (ab1998)
    ICC/IF image of ab1998 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab1998, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

参考文献

This product has been referenced in:

  • Qin QJ  et al. Rhynchophylline ameliorates myocardial ischemia/reperfusion injury through the modulation of mitochondrial mechanisms to mediate myocardial apoptosis. Mol Med Rep 19:2581-2590 (2019). Read more (PubMed: 30720139) »
  • Zhang C  et al. Curcumin induces apoptosis and inhibits angiogenesis in murine malignant mesothelioma. Int J Oncol 53:2531-2541 (2018). Read more (PubMed: 30272283) »
See all 31 Publications for this product

ab1998 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

レビューと Q&A

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1-7 of 7 Abreviews or Q&A

Question

I am interested in looking at AIF as a potential marker of caspase-independent apoptosis. Please can you tell me if this is a reliable marker or if there is another product you would recommend? I don’t see any caspase cleavage but I do see mitochondrial membrane depolarisation and membrane blebbing in my experiments suggesting that some other form of death is occurring.

Also, to use it for this purpose would it be best to look for it in mitochondrial and nuclear fractions by immunoblotting as I presume that FACS & standard immunoblotting would only tell you if it was present in the cell or not or do levels increase after mitochondrial release? Would your cellular fractionation kit ab109719 be suitable for looking at AIF in these fractions?

I am working with rat cells, which of your antibodies is best for this cell type: ab119917, ab32516 or ab1998?

Sorry this is not an area that I am familiar with!

Thanks

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Answer

Thank you for contacting us.

I would suggested searching the potential of this as caspase-independent apoptosis marker in publications. There are many publications related to this protein. Please have a look in pubmed.

As the process triggering apoptosis involves the mitochondrial release of AIF through the mitochondrial membrane, which enters the cytosol, and finally ends up in the cell nucleus where it signals the cell to condense its chromosomes and fragment its DNA molecules in order to prepare for cell death. So the idea would be to compare the AIF level in mitochondrial, cytosolic and nuclear preparations. The kit ab109719 should be able to separate the mitochondrial, cytosolic and nuclear fractions for experiment.

Monoclonal antibodies provide clean WB results so monoclonal will be best suitable e.g. ab119917 and ab32516. This however does not mean polyclonal aren't good, they are good but monoclonal are better.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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https://www.abcam.com/abreviews

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Question

i am encountering a problem with finding the peptide sequences of THE BELOW ON ABCAM WEBSITE ;

Rabbit polyclonal to AIF
ab 1998

Rabbit polyclonal to Bcl2
ab 59348

Rabbit monoclonal [E83-77] to active
Caspase 3
ab 32042

CAN YOU PLS SEND ME THE PEPTIDE SEQUENCES OF THE ABOVE PLS I NEED THEM FOR THE FURTHER STUDIES.

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Answer



ab1998 : https://www.abcam.com/ab1998, where the immunogen is described asa peptide corresponding to amino acids 517 to 531 of human AIF. This sequence is identical tothose of mouse and rat AIF.

ab32042 : https://www.abcam.com/ab32042, where the immunogen is described as : a synthetic peptide corresponding to residues following Ser29 of human Caspase 3 (N terminus of p17 subunit).

ab59348 : https://www.abcam.com/ab59348, where the immunogen is described as a synthesized non phosphopeptide derived from human Bcl2 around the phosphorylation site of threonine 69 (ARTPSP).

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-AIF antibody

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Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Breast cancer tissue and adjacent normal)
Specification
Breast cancer tissue and adjacent normal
Fixative
Formaldehyde
Antigen retrieval step
Other
Permeabilization
No
Blocking step
H2o2 of abcam secondary antibody Kit Expose Kit as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 37°C
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Mrs. Thushara Jayan Pillai

Verified customer

投稿 Oct 24 2011

Western blot abreview for Anti-AIF antibody - Mitochondrial Marker

 Excellent
Abreviews
abreview image
Application
Western blot
Sample
Human Cell lysate - whole cell (SW1573 lung cancer cell line)
Gel Running Conditions
Reduced Denaturing (10% SDS-PAGE)
Loading amount
50 µg
Specification
SW1573 lung cancer cell line
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 8% · Temperature: 25°C
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Abcam user community

Verified customer

投稿 May 27 2011

Question

I´ve been following this problem, even though in this particular Ab I´m not the final user. First of all the concentration the data sheet suggested is the one used from the beginning, also incubation times has been tested ( 1 hour at RT or ON at 4 ºC) so the problem is not that we have a weak band , it is that there is no band at all (even at 4 ºC ON) so we believe your Ab is not working. On the other hand we bought at the same time the COX IV loading control which works fine at the concentration suggested on the data sheet and 1 hour at RT. Increasing Ab concentrations is would be possible if we see a weak signal...remember we have not signal even at ON incubations and it is different to data sheet suggestions and what we expected on this Ab at the time we choose it. More over Carlos thesis work has been done mainly using western blot experiments on same model so I´m clear that we have not problems on the technique.

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Answer

Thank you for your email. In your prior message with the attachment containing the Western blot image, incubation for 1 hr at RT was mentioned but not overnight at 4C. I agree with you and I would expect that you would see some sort of signal with your samples and overnight incubation. As we guarantee our products to work as stated on the online datasheet, I would be happy to offer you a free of charge replacement vial or a credit note or refund. Please let me know how you would like to proceed, and happy holidays!

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Question

BATCH NUMBER 108063 ORDER NUMBER 112828 DESCRIPTION OF THE PROBLEM No signal (No band at all) SAMPLE Purified Rat liver Mitochondria PRIMARY ANTIBODY AB 1998 DETECTION METHOD Super signal west pico (pierce) POSITIVE AND NEGATIVE CONTROLS USED We have tried just rat liver mitochondria lysates using triton x 100, SDS or NP 40. This is my model, and the release of cytochrome c works fine, so I am not interested in other species. ANTIBODY STORAGE CONDITIONS The antibody was Aliquoted upon arrival and store at -30 ?C SAMPLE PREPARATION lysis buffer containing PMSF 1 mM, EDTA 10 mM, Nacl 300 mM, BME 5 mM, NP40 1 %,Tris 50 mM (pH: 7.6) plus protease inhibitors cocktail. AMOUNT OF PROTEIN LOADED 5-30 ?g per lane ELECTROPHORESIS/GEL CONDITIONS Std 12 %SDS-PAGE, run at 170 volts during 1 hour. TRANSFER AND BLOCKING CONDITIONS Wet transfer, 25 volts over night. Blocking solution: 5 % Non fat milk in TBS. 1 Hour RT. SECONDARY ANTIBODY Goat anti-Rabbit- HRP conjugated (AP 132P, Chemicon) HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 4 HAVE YOU RUN A "NO PRIMARY" CONTROL? No WHAT STEPS HAVE YOU ALTERED? NONE ADDITIONAL NOTES I have experience with several anti-bodies. For most of them the western blot condition are the same with not troubles of any kind. I am investigating the release of apoptotic factors from isolated mitochondria induced by Bax. The detection in this experiments of cytochrome c works fine. But now I wanted to try AIF release in this model. But the AB 1998 anti-body did not work.

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Answer

Thank you for your enquiry and I'm sorry to hear that you are experiencing difficulty with ab1998. At this time I would like to make the following suggestions in order to help improve your results. I would suggest extending the incubation period with the primary antibody to overnight at 4C and also increasing the concentration of ab1998 to 1:500. I also suggest running a positive control - K562 cell lysate is recommended for ab1998. Please contact us again if you have any additional questions, and happy holidays.

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Question

Hello! I would like use the rabbit polyclonal AIF antibody (ab1998 or ab2086) on paraffin sections 5-8 micron. Do you have some experience on this possibility?; can you give me some suggestions?

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Answer

This antibody has not been tested for IHC, therefore we cannot guarantee results.

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