Recombinant Anti-ALIX antibody [EPR23653-32] (ab275377)
![Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-467652-anti-alix-antibody-epr23653-32-western-blot-wildtype-hek293-pdcd6ip-knockout-hek293-hela-human-spleen.jpg "Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23653-32] to ALIX
- Suitable for: WB, ICC/IF, Flow Cyt (Intra), IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-ALIX antibody [EPR23653-32]
See all ALIX primary antibodies -
Description
Rabbit monoclonal [EPR23653-32] to ALIX -
Host species
Rabbit -
Tested applications
Suitable for: WB, ICC/IF, Flow Cyt (Intra), IPmore details
Unsuitable for: IHC-P -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: C6, RAW 264.7, PC-12, NIH/3T3, K-562, HEK-293, HeLa, HCT116, MCF7 and Jurkat whole cell lysates; Human brain tissue lysate; Mouse brain tissue lysate; Rat brain tissue lysate. ICC/IF: NIH/3T3 and HeLa cells. Flow Cyt (intra): NIH/3T3 and HeLa cells. IP: K562 whole cell lysate.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23653-32 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab275377 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 |
|---|---|---|
| WB |
1/1000. Detects a band of approximately 80, 90, 100 kDa (predicted molecular weight: 96 kDa).
|
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| ICC/IF |
1/5000.
|
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| Flow Cyt (Intra) |
1/500.
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| IP |
1/30.
|
| Notes |
|---|
|
WB
1/1000. Detects a band of approximately 80, 90, 100 kDa (predicted molecular weight: 96 kDa). |
|
ICC/IF
1/5000. |
|
Flow Cyt (Intra)
1/500. |
|
IP
1/30. |
Target
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Function
Class E VPS protein involved in concentration and sorting of cargo proteins of the multivesicular body (MVB) for incorporation into intralumenal vesicles (ILVs) that are generated by invagination and scission from the limiting membrane of the endosome. Binds to the phospholipid lysobisphosphatidic acid (LBPA) which is abundant in MVBs internal membranes. The MVB pathway appears to require the sequential function of ESCRT-O, -I,-II and -III complexes. The ESCRT machinery also functions in topologically equivalent membrane fission events, such as the terminal stages of cytokinesis and enveloped virus budding (HIV-1 and other lentiviruses). Appears to be an adapter for a subset of ESCRT-III proteins, such as CHMP4, to function at distinct membranes. Required for completion of cytokinesis. Involved in HIV-1 virus budding. Can replace TSG101 it its role of supporting HIV-1 release; this function implies the interaction with CHMP4B. May play a role in the regulation of both apoptosis and cell proliferation. -
Sequence similarities
Contains 1 BRO1 domain. -
Cellular localization
Cytoplasm > cytosol. Melanosome. Cytoplasm > cytoskeleton > centrosome. Identified by mass spectrometry in melanosome fractions from stage I to stage IV. Colocalized with CEP55 in the midbody during cytokinesis. Colocalized with CEP55 at centrosomes of non-dividing cells. - Information by UniProt
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Database links
- Entrez Gene: 10015 Human
- Entrez Gene: 18571 Mouse
- Entrez Gene: 501083 Rat
- Omim: 608074 Human
- SwissProt: Q8WUM4 Human
- SwissProt: Q9WU78 Mouse
- SwissProt: Q9QZA2 Rat
- Unigene: 475896 Human
see all -
Alternative names
- AIP1 antibody
- ALG 2 interacting protein 1 antibody
- ALG-2-interacting protein 1 antibody
see all
Images
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Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)All lanes : Anti-ALIX antibody [EPR23653-32] (ab275377) at 1/1000 dilution
Lane 1 : Wild-type HEK-293 cell lysate
Lane 2 : PDCD6IP knockout HEK-293 cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : Human Spleen tissue lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 96 kDa
Observed band size: 96 kDaLanes 1 - 4: Merged signal (red and green). Green - ab275377 observed at 96 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
ab275377 was shown to react with PDCD6IP in wild-type HEK-293T cells in Western blot with loss of signal observed in PDCD6IP knockout cell line ab260864 (PDCD6IP knockout cell lysate ab261656). Wild-type HEK-293T and PDCD6IP knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5 % milk in TBS-T (0.1 % Tween®) before incubation with ab275377 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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![Immunocytochemistry/ Immunofluorescence - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-2-anti-alix-antibody-epr2365332-immunocytochemistry-hela.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-ALIX antibody [EPR23653-32] (ab275377)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labelling ALIX with ab275377 at 1/5000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488, ab150077) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HeLa cells is observed. Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594, ab195889) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488, ab150077) at 1/1000 dilution.
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![Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-409473-anti-alix-antibody-epr2365332-western-blot-lysate1.jpg)
Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)All lanes : Anti-ALIX antibody [EPR23653-32] (ab275377) at 1/1000 dilution
Lane 1 : Mouse brain tissue lysate
Lane 2 : Rat brain tissue lysate
Lane 3 : C6 (rat glial tumor glial cell) whole cell lysate
Lane 4 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 5 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate
Lane 6 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution
Predicted band size: 96 kDa
Observed band size: 80/90/100 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
The expression pattern is consistent with what has been described in the literature (PMID: 24834918, 26935291, 28322231).
Exposure time: Lane1-2: 10 seconds Lane3-6: 8 seconds.
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![Flow Cytometry (Intracellular) - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-3-anti-alix-antibody-epr2365332-flow-cytometry-hela.jpg)
Flow Cytometry (Intracellular) - Anti-ALIX antibody [EPR23653-32] (ab275377)Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell line) cells labelling ALIX with ab275377 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).
A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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![Immunoprecipitation - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-5-anti-alix-antibody-epr2365332-immunoprecipitation-k562.jpg)
Immunoprecipitation - Anti-ALIX antibody [EPR23653-32] (ab275377)ALIX was immunoprecipitated from 0.35 mg K-562 (human chronic myelogenous leukemia lymphoblast cell line) whole cell lysate with ab275377 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab275377 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: K-562 whole cell lysate 10 ug
Lane 2: ab275377 IP in K-562 whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab275377 in K-562 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3 seconds
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![Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-409472-anti-alix-antibody-epr2365332-western-blot-lysate.jpg)
Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)Anti-ALIX antibody [EPR23653-32] (ab275377) at 1/1000 dilution + Human brain tissue lysate at 20 µg
Secondary
VeriBlot for IP secondary antibody(HRP)(ab131366) at 1/1000 dilution
Predicted band size: 96 kDa
Observed band size: 80/90/100 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
The expression pattern is consistent with what has been described in the literature (PMID: 24834918, 26935291, 28322231).
Exposure time: 10 seconds.
-
![Flow Cytometry (Intracellular) - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-4-anti-alix-antibody-epr2365332-flow-cytometry-nih3t3.jpg)
Flow Cytometry (Intracellular) - Anti-ALIX antibody [EPR23653-32] (ab275377)Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized NIH/3T3 (mouse embryonic fibroblast cell line) cells labelling ALIX with ab275377 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).
A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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![Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-409471-anti-alix-antibody-epr2365332-western-blot-myelogenous.jpg)
Western blot - Anti-ALIX antibody [EPR23653-32] (ab275377)All lanes : Anti-ALIX antibody [EPR23653-32] (ab275377) at 1/1000 dilution
Lane 1 : K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate
Lane 2 : HEK-293 (human embryonic kidney epithelial cell) whole cell lysate
Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 4 : HCT116 (human colorectal carcinoma epithelial cell) whole cell lysate
Lane 5 : MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate
Lane 6 : Jurkat (human T cell leukemia T lymphocyte) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution
Predicted band size: 96 kDa
Observed band size: 90/100 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
The expression pattern is consistent with what has been described in the literature (PMID: 24834918, 26935291, 28322231).
Exposure time: 10 seconds.
-
![Immunocytochemistry/ Immunofluorescence - Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-1-anti-alix-antibody-epr2365332-immunocytochemistry-nih3t3.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-ALIX antibody [EPR23653-32] (ab275377)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embyro fibroblast cell line) cells labelling ALIX with ab275377 at 1/5000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488, ab150077 ) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in NIH/3T3 cells. Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594, ab195889) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
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![Anti-ALIX antibody [EPR23653-32] (ab275377)](https://www.abcam.com/ps/products/275/ab275377/Images/ab275377-9-benefits-of-recombinant-antibodies.png)
Anti-ALIX antibody [EPR23653-32] (ab275377)
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
Certificate of Compliance
References (1)
ab275377 has been referenced in 1 publication.
- Zhang R et al. Exosomal SNHG16 secreted by CSCs promotes glioma development via TLR7. Stem Cell Res Ther 12:349 (2021). PubMed: 34134771