Recombinant Anti-Optineurin antibody [EPR20654] KO Tested (ab213556)

Product name

Anti-Optineurin antibody [EPR20654]
See all Optineurin primary antibodies
Description

Rabbit monoclonal [EPR20654] to Optineurin
Host species

Rabbit
Tested applications

Suitable for: IP, IHC-P, WB, IHC-Frmore details
Unsuitable for: ICC/IF
Species reactivity

Reacts with: Mouse, Rat, Human
Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Human fetal brain, fetal kidney, placenta and skeletal muscle lysates; Rat brain, retina, placenta and heart lysates; Mouse retina and placenta lysates; 293T, U-2 OS and NIH/3T3 whole cell lysates. IHC-P: Human retina and cerebrum tissues; Mouse retina and cerebrum tissues; Rat cerebellum tissue. IP: 293T and U-2 OS whole cell lysate.
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

EPR20654
Isotype

IgG
Research areas

Alternative Versions
- Anti-Optineurin antibody [EPR20654] - BSA and Azide free (ab240979)
Compatible Secondaries
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Recombinant Protein
- Recombinant Human Optineurin protein (His tag) (ab235021)
Related Products
- VeriBlot for IP Detection Reagent (HRP) (ab131366)
- Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) (ab93684)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab213556 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
IP		1/40.
IHC-P		1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB		1/1000. Detects a band of approximately 68 kDa (predicted molecular weight: 66 kDa).
IHC-Fr		1/100.

Notes
IP 1/40.
IHC-P 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB 1/1000. Detects a band of approximately 68 kDa (predicted molecular weight: 66 kDa).
IHC-Fr 1/100.

Application notes

Is unsuitable for ICC/IF.

Function

Plays an important role in the maintenance of the Golgi complex, in membrane trafficking, in exocytosis, through its interaction with myosin VI and Rab8. Links myosin VI to the Golgi complex and plays an important role in Golgi ribbon formation. Negatively regulates the induction of IFNB in response to RNA virus infection. Plays a neuroprotective role in the eye and optic nerve. Probably part of the TNF-alpha signaling pathway that can shift the equilibrium toward induction of cell death. May act by regulating membrane trafficking and cellular morphogenesis via a complex that contains Rab8 and hungtingtin (HD). May constitute a cellular target for adenovirus E3 14.7, an inhibitor of TNF-alpha functions, thereby affecting cell death.
Tissue specificity

Present in acqueous humor of the eye (at protein level). Highly expressed in trabecular meshwork. Expressed nonpigmented ciliary epithelium, retina, brain, adrenal cortex, fetus, lymphocyte, fibroblast, skeletal muscle, heart, liver, brain and placenta.
Involvement in disease

Defects in OPTN are the cause of primary open angle glaucoma type 1E (GLC1E) [MIM:137760]. Primary open angle glaucoma (POAG) is characterized by a specific pattern of optic nerve and visual field defects. The angle of the anterior chamber of the eye is open, and usually the intraocular pressure is increased. The disease is asymptomatic until the late stages, by which time significant and irreversible optic nerve damage has already taken place.
Defects in OPTN are a cause of susceptibility to normal pressure glaucoma (NPG) [MIM:606657].
Defects in OPTN are the cause of amyotrophic lateral sclerosis type 12 (ALS12) [MIM:613435]. It is a neurodegenerative disorder affecting upper motor neurons in the brain and lower motor neurons in the brain stem and spinal cord, resulting in fatal paralysis. Sensory abnormalities are absent. Death usually occurs within 2 to 5 years. The etiology of amyotrophic lateral sclerosis is likely to be multifactorial, involving both genetic and environmental factors. The disease is inherited in 5-10% of the cases.
Domain

Ubiquitin-binding motif (UBAN) is essential for its inhibitory function, subcellular localization and interaction with TBK1.
Post-translational
modifications

Phosphorylated. Phosphorylation is induced by phorbol esters and decreases its half-time.
Cellular localization

Cytoplasm > perinuclear region. Golgi apparatus. Golgi apparatus > trans-Golgi network. Found in the perinuclear region and associates with the Golgi apparatus. Colocalizes with MYO6 and RAB8 at the Golgi complex and in vesicular structures close to the plasma membrane.
Target information above from: UniProt accession Q96CV9 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 10133 Human
- Entrez Gene: 71648 Mouse
- Entrez Gene: 246294 Rat
- Omim: 602432 Human
- SwissProt: Q96CV9 Human
- SwissProt: Q8K3K8 Mouse
- SwissProt: Q8R5M4 Rat
- Unigene: 332706 Human
- Unigene: 116942 Mouse
- Unigene: 63435 Rat
see all
Alternative names
- 14.7K interacting protein antibody
- Ag9 C5 antibody
- ALS12 antibody
- E3 14.7K interacting protein antibody
- E3-14.7K-interacting protein antibody
- FIP 2 antibody
- FIP-2 antibody
- FIP2 antibody
- Glaucoma 1 open angle E (adult onset) antibody
- Glaucoma 1 open angle E antibody
- GLC1E antibody
- HIP 7 antibody
- HIP-7 antibody
- HIP7 antibody
- Huntingtin interacting protein 7 antibody
- Huntingtin interacting protein HYPL antibody
- Huntingtin interacting protein L antibody
- Huntingtin yeast partner L antibody
- Huntingtin-interacting protein 7 antibody
- Huntingtin-interacting protein L antibody
- HYPL antibody
- Injury inducible protein I 55 antibody
- NEMO related protein antibody
- NEMO-related protein antibody
- NRP antibody
- Optic neuropathy inducing protein antibody
- Optic neuropathy-inducing protein antibody
- Optineurin antibody
- OPTN antibody
- OPTN_HUMAN antibody
- TFIIIA IntP antibody
- TFIIIA-IntP antibody
- Transcription factor IIIA interacting protein antibody
- Transcription factor IIIA-interacting protein antibody
- Tumor necrosis factor alpha inducible cellular protein containing leucine zipper domains antibody
see all

Immunohistochemistry (Frozen sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

ab213556 staining Optineurin in Mouse retina tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with 4% paraformaldehyde, permeabilized with 0.2% Triton. Samples were incubated with primary antibody (1/100). An Alexa Fluor^® 488 Goat anti-Rabbit secondary (1/1000) was used as the secondary antibody. Counter stained with DAPI.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20)

Positive staining on mouse retina (PMID: 15607428).
Western blot - Anti-Optineurin antibody [EPR20654] (ab213556)

All lanes : Anti-Optineurin antibody [EPR20654] (ab213556) at 1/5000 dilution

Lane 1 : Wild-type U-2 OS cell lysate
Lane 2 : OPTN knockout U-2 OS cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 66 kDa

ab213556 was shown to react with OPTN in wild-type U-2 OS cells in Western blot with loss of signal observed in a OPTN knockout cell line. Wild-type U-2 OS and OPTN knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab213556 overnight at 4 °C at a 1/5000 dilution. Blots were incubated with goat anti-rabbit HRP secondary antibodies at 1/5000 before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunohistochemical analysis of paraffin-embedded human retina tissue labeling Optineurin with ab213556 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on human retina (PMID: 15607428).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunoprecipitation - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunoprecipitation of OPTN in c-43 cells. Lysates were prepared and immunoprecipitation was performed using 1.0 μg of ab213556 pre-coupled to prot.A-Sepharose beads. Samples were washed and processed for western blot with ab213556 at 1/5000. This data was kindly provided by the YCharOS Inc., an open science company with the mission of characterizing every commercially available antibody reagent. Abcam are working with YCharOS to support their mission of antibody characterisation using knock out cell lines.
Western blot - Anti-Optineurin antibody [EPR20654] (ab213556)

All lanes : Anti-Optineurin antibody [EPR20654] (ab213556) at 1/1000 dilution

Lane 1 : Human skeletal muscle lysate at 20 µg
Lane 2 : 293T (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg
Lane 3 : Rat brain lysate at 20 µg
Lane 4 : Mouse retina lysate at 20 µg
Lane 5 : Rat retina lysate at 20 µg
Lane 6 : Rat placenta lysate at 20 µg
Lane 7 : NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 µg

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 66 kDa
Observed band size: 68 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1-3/7: 3 minutes; Lane 4-6: 10 seconds.

The MW observed is consistent with the literature (PMID 20174559, PMID 24983867, PMID 11834836).
Immunoprecipitation - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunoprecipitation of OPTN in U-2 OS cells. Lysates were prepared and immunoprecipitation was performed using 1.0 μg of ab213556 pre-coupled to prot.A-Sepharose beads. Samples were washed and processed for western blot with ab213556 at 1/5000. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitate. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
Immunohistochemistry (Frozen sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

ab213556 staining Optineurin in Rat retina tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with 4% paraformaldehyde, permeabilized with 0.2% Triton. Samples were incubated with primary antibody (1/100). An Alexa Fluor^® 488 Goat anti-Rabbit secondary (1/1000) was used as the secondary antibody. Counter stained with DAPI.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20)

Positive staining on rat retina (PMID: 15607428).
Western blot - Anti-Optineurin antibody [EPR20654] (ab213556)

All lanes : Anti-Optineurin antibody [EPR20654] (ab213556) at 1/1000 dilution

Lane 1 : Human fetal brain lysate
Lane 2 : Human fetal kidney lysate
Lane 3 : Human placenta lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution

Predicted band size: 66 kDa
Observed band size: 68 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1: 3 seconds; Lane 2/3: 30 seconds.

The MW observed is consistent with the literature (PMID 20174559, PMID 24983867, PMID 11834836).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labeling Optineurin with ab213556 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on human cerebrum (PMID: 26303227).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Optineurin antibody [EPR20654] (ab213556)

All lanes : Anti-Optineurin antibody [EPR20654] (ab213556) at 1/1000 dilution

Lane 1 : Rat heart lysate
Lane 2 : Mouse placenta lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 66 kDa
Observed band size: 68 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1: 3 minutes; Lane 2: 10 seconds.

The MW observed is consistent with the literature (PMID 20174559, PMID 24983867, PMID 11834836).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunohistochemical analysis of paraffin-embedded mouse retina tissue labeling Optineurin with ab213556 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on mouse retina (PMID: 15607428).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labeling Optineurin with ab213556 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on mouse cerebrum (PMID: 26303227).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR20654] (ab213556)

Immunohistochemical analysis of paraffin-embedded rat cerebellum tissue labeling Optineurin with ab213556 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on rat cerebellum (PMID: 26303227).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunoprecipitation - Anti-Optineurin antibody [EPR20654] (ab213556)

Optineurin was immunoprecipitated from 0.35 mg of 293T (Human epithelial cell line from embryonic kidney) whole cell lysate with ab213556 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab213556 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution

Lane 1: 293T whole cell lysate 10 μg (Input).

Lane 2: ab213556 IP in 293T whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab213556 in 293T whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 3 minutes.
Anti-Optineurin antibody [EPR20654] (ab213556)

Western blot protocols
Immunoprecipitation protocols
Immunohistochemistry protocols

Click here to view the general protocols

Datasheet download

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Publishing research using ab213556? Please let us know so that we can cite the reference in this datasheet.

ab213556 has been referenced in 1 publication.

Hu S et al. Structure of Myosin VI/Tom1 complex reveals a cargo recognition mode of Myosin VI for tethering. Nat Commun 10:3459 (2019). PubMed: 31371777

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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

Recombinant Protein

Related Products

Applications

The Abpromise guarantee

Target

Function

Tissue specificity

Involvement in disease

Domain

Post-translationalmodifications

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

Datasheet download

Certificate of Compliance

References (1)

Customer reviews and Q&As

Post-translational
modifications