Recombinant Anti-UBE3A antibody [EPR23077-14] (ab272168)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23077-14] to UBE3A
- Suitable for: Indirect ELISA, IP, WB, ICC/IF, Flow Cyt (Intra)
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
-
Product name
Anti-UBE3A antibody [EPR23077-14]
See all UBE3A primary antibodies -
Description
Rabbit monoclonal [EPR23077-14] to UBE3A -
Host species
Rabbit -
Tested applications
Suitable for: Indirect ELISA, IP, WB, ICC/IF, Flow Cyt (Intra)more 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.
-
Positive control
- WB: HeLa, K562, HepG2,A549, HEK-293T, PC-12, PC-12 (treated with 10 uM MG-132 for 4 hours), RAW 264.7 and RAW 264.7 (treated with 10 uM MG-132 for 4 hours) whole cell lysates; Mouse spleen tissue lysate;, Rat brain tissue lysate. ICC/IF: HeLa and RAW 264.7 cells. Flow Cyt (intra): HeLa and RAW 264.7 cells. IP: K562 and RAW 264.7 whole cell lysates.
-
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
-
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
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
-
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23077-14 -
Isotype
IgG -
Research areas
Associated products
-
Alternative Versions
-
Compatible Secondaries
-
Isotype control
-
Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab272168 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 |
---|---|---|
Indirect ELISA |
Use a concentration of 1 µg/ml.
|
|
IP |
1/30.
|
|
WB |
1/1000. Detects a band of approximately 100, 37 kDa (predicted molecular weight: 100 kDa).
|
|
ICC/IF |
1/100.
|
|
Flow Cyt (Intra) |
1/50.
|
Notes |
---|
Indirect ELISA
Use a concentration of 1 µg/ml. |
IP
1/30. |
WB
1/1000. Detects a band of approximately 100, 37 kDa (predicted molecular weight: 100 kDa). |
ICC/IF
1/100. |
Flow Cyt (Intra)
1/50. |
Target
-
Function
E3 ubiquitin-protein ligase which accepts ubiquitin from an E2 ubiquitin-conjugating enzyme in the form of a thioester and transfers it to its substrates. Several substrates have been identified including the RAD23A and RAD23B, MCM7 (which is involved in DNA replication), annexin A1, the PML tumor suppressor, and the cell cycle regulator CDKN1B. Catalyzes the high-risk human papilloma virus E6-mediated ubiquitination of p53/TP53, contributing to the neoplastic progression of cells infected by these viruses. Additionally, may function as a cellular quality control ubiquitin ligase by helping the degradation of the cytoplasmic misfolded proteins. Finally, UBE3A also promotes its own degradation in vivo. Plays an important role in the regulation of the circadian clock: involved in the ubiquitination of the core clock component ARNTL/BMAL1, leading to its proteasomal degradation (PubMed:24728990). -
Pathway
Protein modification; protein ubiquitination. -
Involvement in disease
Angelman syndrome -
Sequence similarities
Contains 1 HECT (E6AP-type E3 ubiquitin-protein ligase) domain. -
Post-translational
modificationsPhosphorylation at Tyr-659 by ABL1 impairs E3 ligase activity and protects p53/TP53 from degradation in (HPV)-infected cells. -
Cellular localization
Nucleus. Cytoplasm. - Information by UniProt
-
Database links
- Entrez Gene: 7337 Human
- Entrez Gene: 22215 Mouse
- Entrez Gene: 361585 Rat
- Omim: 601623 Human
- SwissProt: Q05086 Human
- SwissProt: O08759 Mouse
- Unigene: 598862 Human
- Unigene: 9002 Mouse
-
Alternative names
- ANCR antibody
- Angelman syndrome antibody
- AS antibody
see all
Images
-
All lanes : Anti-UBE3A antibody [EPR23077-14] (ab272168) at 1/1000 dilution
Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) treated with 10 uM MG-132 for 24 hours, whole cell lysate
Lane 2 : HEK-293T (human embryonic kidney epithelial cell) 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: 100 kDa
Observed band size: 100 kDaBlocking and diluting buffer and concentration: 5% NFDM/TBST.
Fresh lysate was used in lane 2.
Exposure time: Lane 1: 3 minutes; Lane 2: 48 seconds.
-
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labelling UBE3A with ab272168 at 1/100 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear and weak cytoplasmic staining in HeLa cell line ab195889. Anti-alpha Tubulin antibody (Alexa Fluor® 594) 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 1000 dilution.
-
Indirect ELISA using ab272168 at varying antibody concentrations (1000-0 ng/ml) and Human UBE3A antigen at 1000 ng/ml. Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as a secondary antibody.
-
Intracellular flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol-permeabilized HeLa (Human cervix adenocarcinoma epithelial cell) cells labelling UBE3A with ab272168 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control 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.
-
UBE3A was immunoprecipitated from 0.35 mg K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate with ab272168 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab272168 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate 10 ug
Lane 2: ab272168 IP in K-562 whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab272168 in K-562 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 15 seconds
A 37 kDa degraded band is observed.
-
All lanes : Anti-UBE3A antibody [EPR23077-14] (ab272168) at 1/1000 dilution
Lane 1 : PC-12 (rat adrenal gland pheochromocytoma ) whole cell lysate
Lane 2 : PC-12 treated with 10 µM MG-132 for 4 hours, whole cell lysate
Lane 3 : RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 4 : RAW 264.7 treated with 10 µM MG-132 for 4 hours, whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 100 kDa
Observed band size: 100,37 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.
Generation of the 37-kDa degraded fragment can be inhibited/reduced by MG-132 treatment (lanes 2 and 4).
-
All lanes : Anti-UBE3A antibody [EPR23077-14] (ab272168) at 1/1000 dilution
Lane 1 : Mouse spleen tissue lysate
Lane 2 : Rat brain tissue lysate
Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 4 : K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate
Lane 5 : HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 6 : HEK-293T (human embryonic kidney epithelial cell) whole cell lysate
Lane 7 : A549 (human lung carcinoma epithelial cell) 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: 100 kDa
Observed band size: 100,37 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
A 37 kDa degraded band is observed. MG132 treatment or freshly made lysates can decrease the degradation.
Exposure time: 3 minutes
-
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized RAW 264.7 cells labelling UBE3A with ab272168 at 1/100 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear and weak cytoplasmic staining in RAW 264.7 cell line. ab195889 Anti-alpha Tubulin antibody (Alexa Fluor® 594) 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 1000 dilution.
-
Intracellular flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol-permeabilized RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) cells labelling UBE3A with ab272168 at 1/50 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control 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.
-
UBE3A was immunoprecipitated from 0.35 mg RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) (treated with 10 μM MG-132 for 4 hours) whole cell lysate with ab272168 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab272168 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) treated with 10 μM MG-132 for 4 hours, whole cell lysate 10 ug
Lane 2: ab272168 IP in RAW 264.7 treated with 10 μM MG-132 for 4 hours, whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab272168 in RAW 264.7 treated with 10 μM MG-132 for 4 hours, whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 10 seconds
A 37 kDa degraded band is observed.
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
-
SDS download
-
Datasheet download
Certificate of Compliance
References (0)
ab272168 has not yet been referenced specifically in any publications.