Anti-AIF antibody [E20] - Mitochondrial Marker (ab32516)

Overview

  • Product nameAnti-AIF antibody [E20] - Mitochondrial Marker
    See all AIF primary antibodies
  • Description
    Rabbit monoclonal [E20] to AIF - Mitochondrial Marker
  • Tested applicationsSuitable for: ICC/IF, WB, IHC-P, Flow Cyt, IP, IHC-Frmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    A synthetic peptide correspndonding to residues near the C terminus of Human AIF.

  • Positive control
    • WB: K562 cell lysate. IHC-P: Human cervical carcinoma tissue.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

    Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

     

    Alternative versions available:

    Anti-AIF antibody - Mitochondrial Marker (Alexa Fluor® 647) [E20] (ab196847)

    Anti-AIF antibody - Mitochondrial Marker (Biotin) [E20] (ab197526)

Properties

Applications

Our Abpromise guarantee covers the use of ab32516 in the following tested applications.

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

Application Abreviews Notes
ICC/IF 1/500.
WB 1/1000. Detects a band of approximately 67 kDa (predicted molecular weight: 67 kDa).
IHC-P Use at an assay dependent concentration.
Flow Cyt 1/50.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IP Use at an assay dependent concentration.
IHC-Fr Use at an assay dependent concentration. PubMed: 23118224

Target

  • FunctionProbable 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.
  • Involvement in diseaseDefects 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.
  • Sequence similaritiesBelongs to the FAD-dependent oxidoreductase family.
  • Post-translational
    modifications
    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.
  • Cellular localizationMitochondrion 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
  • Database links
  • Alternative names
    • 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

Anti-AIF antibody [E20] - Mitochondrial Marker images

  • Anti-AIF antibody [E20] - Mitochondrial Marker (ab32516) at 1/1000 dilution + K562 cell lysate

    Predicted band size : 67 kDa
    Observed band size : 67 kDa
  • AIF was immunoprecipitated using 0.5mg K562 whole cell extract, 5µg of Rabbit monoclonal to AIF and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, K562 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab32516.
    Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
    Band: 67kDa: AIF
  • Ab32516, at a 1/500 dilution, staining AIF in paraffin embedded human cervical carcinoma tissue by Immunohistochemistry.
  • ICC/IF image of ab32516 stained HeLa cells. The cells were 4% PFA 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 (ab32516, 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.

  • Overlay histogram showing K562 cells stained with ab32516 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab32516, 1/50 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in K562 cells fixed with methanol (5 min)/permeabilized with 0.1% PBS-Tween used under the same conditions.

References for Anti-AIF antibody [E20] - Mitochondrial Marker (ab32516)

This product has been referenced in:
  • Fan WY  et al. The cytotoxic effect of oxybuprocaine on human corneal epithelial cells by inducing cell cycle arrest and mitochondria-dependent apoptosis. Hum Exp Toxicol N/A:N/A (2016). Read more (PubMed: 27590991) »
  • Tang Y  et al. Quantitative proteomic analysis of HER2 normal and overexpressing MCF-7 breast cancer cells revealed proteomic changes accompanied with HER2 gene amplification. J Proteomics 91C:200-209 (2013). Human . Read more (PubMed: 23851309) »

See all 10 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Human Tissue sections (skin)
Specification skin
Fixative Paraformaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: 100mM sodium cytrate
Blocking step BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
Username

Ms. Hadas Pahima

Verified customer

Submitted Oct 11 2012

Thank you very much for contacting us and for your feedback.

We have published this feedback in the scientific support section of the datasheet and it is now available to all the scientific community. Next, we would like to encourage you to pu...

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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 ...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Tissue lysate - whole (brain)
Loading amount 30 µg
Specification brain
Gel Running Conditions Reduced Denaturing (10)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Username

ilan sela

Verified customer

Submitted Aug 09 2012

Vielen Dank für Ihre Antwort. Ich konnte anhand Ihrer Angaben die wahrscheinlichste Charge ausfindig machen: Die Konzentration Ihrer Charge ist xxxx. Ich möchte Sie darauf hinweisen, dass diese scheinbar niedrige Konzentration völlig ausreichend ist. D...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Rat Tissue lysate - whole (Skeletal Muscle)
Loading amount 30 µg
Specification Skeletal Muscle
Gel Running Conditions Reduced Denaturing (10%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 0.5% · Temperature: RT°C
Username

Abcam user community

Verified customer

Submitted May 15 2009

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