Human AIFM1 (AIF) knockout HEK-293T cell lysate (ab256834)
Overview
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Product name
Human AIFM1 (AIF) knockout HEK-293T cell lysate
See all AIF kits -
Product overview
Knockout cell lysate achieved by CRISPR/Cas9. -
Parental Cell Line
HEK293T -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 2 and 23 bp deletion in exon 2 and 2 bp insertion in exon 2. -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Reconstitution notes
To use as WB control, resuspend the lyophilizate in 50 µL of LDS* Sample Buffer to have a final concentration of 2 mg/ml. For reducing conditions, we recommend a final concentration of 0.1 M DTT.*Usage of SDS sample buffer is not recommended with these lyophilized lysates.
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Notes
Lysate preparation: Our lysates are made using RIPA buffer to which we add a protease inhibitor cocktail and phosphatase inhibitor cocktail (ratio: 300:100:10). This means that the protein of interest is denatured. If you require a native form of the protein please use the live cell version - found here. Please refer to our lysis protocol for further details on how our lysates are prepared.
User storage instructions: Lyophilizate may be stored at 4°C. After reconstitution, store at -20°C for short-term storage or -80°C for long-term storage.
Access thousands of knockout cell lysates, generated from commonly used cancer cell lines.
See here for more information on knockout cell lysates.Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
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Tested applications
Suitable for: WBmore details
Properties
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Storage instructions
Store at -80°C. Please refer to protocols. -
Components 1 kit ab260178 - Human AIFM1 knockout HEK293T cell lysate 1 x 100µg ab255553 - Human wild-type HEK293T cell lysate 1 x 100µg -
Research areas
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Cell type
epithelial -
STR Analysis
Amelogenin X D5S818: 8, 9 D13S317: 12, 14 D7S820: 11 D16S539: 9, 13 vWA: 16, 19 TH01: 7, 9.3 TPOX: 11 CSF1PO: 11, 12
Target
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Function
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. -
Involvement in disease
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. -
Sequence similarities
Belongs to the FAD-dependent oxidoreductase family. -
Post-translational
modificationsUnder 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 localization
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
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Alternative names
- AIFM1
- AIFM1_HUMAN
- Apoptosis inducing factor
see all
Associated products
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KO cell lines
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab256834 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 |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 67 kDa.
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Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 67 kDa. |
Images
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Lane 1: Wild-type HEK-293T cell lysate (20µg)
Lane 2: AIFM1 knockout HEK-293T cell lysate (20µg)
Lanes 1- 2: Merged signal (red and green). Green - ab32516 observed at 67 kDa. Red - loading control ab8245 observed at 37 kDa.
ab32516 Anti-AIF antibody [E20] - Mitochondrial Marker was shown to specifically react with AIF in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266347 (knockout cell lysate ab256834) was used. Wild-type and AIF knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32516 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging. -
Allele-1: 23 bp deletion in exon 2
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Allele-2: 1 bp insertion in exon 2
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Allele-3: 2 bp insertion in exon 2
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab256834 has not yet been referenced specifically in any publications.