Recombinant Anti-UBE3A antibody [EPR23077-14] (ab272168)

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Western blot - 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

Carrier Free

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
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

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
- Neuroscience
- Processes

Alternative Versions
- Anti-UBE3A antibody [EPR23077-14] - BSA and Azide free (ab272173)
Compatible Secondaries
- Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
- Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Related Products
- Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (ab195889)

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.

Application notes

Is unsuitable for IHC-P.

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
modifications

Phosphorylation at Tyr-659 by ABL1 impairs E3 ligase activity and protects p53/TP53 from degradation in (HPV)-infected cells.
Cellular localization

Nucleus. Cytoplasm.
Target information above from: UniProt accession Q05086 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

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
- CTCL tumor antigen se37 2 antibody
- CTCL tumor antigen se37-2 antibody
- E6 AP antibody
- E6AP antibody
- E6AP ubiquitin protein ligase antibody
- E6AP ubiquitin-protein ligase antibody
- EPVE6AP antibody
- FLJ26981 antibody
- HECT-type ubiquitin transferase E3A antibody
- HPVE6A antibody
- Human papilloma virus E6 associated protein Angelman syndrome antibody
- Human papilloma virus E6 associated protein antibody
- Human papillomavirus E6-associated protein antibody
- NY REN 54 antigen antibody
- Oncogenic protein associated protein E6 AP antibody
- Oncogenic protein associated protein E6AP antibody
- Oncogenic protein-associated protein E6-AP antibody
- Renal carcinoma antigen NY REN 54 antibody
- Renal carcinoma antigen NY-REN-54 antibody
- UBE 3A antibody
- Ube3a antibody
- UBE3A protein antibody
- UBE3A_HUMAN antibody
- Ubiquitin protein ligase E3A antibody
- Ubiquitin-protein ligase E3A antibody
see all

Western blot - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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 kDa

Blocking 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.
Immunocytochemistry/ Immunofluorescence - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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 - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Flow Cytometry (Intracellular) - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Immunoprecipitation - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Western blot - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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).
Western blot - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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
Immunocytochemistry/ Immunofluorescence - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Flow Cytometry (Intracellular) - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Immunoprecipitation - Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.
Anti-UBE3A antibody [EPR23077-14] (ab272168)

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.

Click here to view the general protocols

SDS download

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Publishing research using ab272168? Please let us know so that we can cite the reference in this datasheet.

ab272168 has not yet been referenced specifically in any publications.

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Key features and details

Related conjugates and formulations

Overview

Product name

Description

Host species

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Isotype control

Related Products

Applications

The Abpromise guarantee

Target

Function

Pathway

Involvement in disease

Sequence similarities

Post-translationalmodifications

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

References (0)

Customer reviews and Q&As

Post-translational
modifications