Overview

  • Product name
    Anti-Parkin antibody [PRK8]
    See all Parkin primary antibodies
  • Description
    Mouse monoclonal [PRK8] to Parkin
  • Host species
    Mouse
  • Tested applications
    Suitable for: IHC-Fr, IHC-P, Flow Cyt, WB, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Drosophila melanogaster
  • Immunogen

    Recombinant full length Human Parkin

  • Epitope
    The epitope is the second ring domain (aa 399-465).
  • Positive control
    • WB: SHSY-5Y whole cell lysate and in human, mouse and rat brain tissue lysates. Flow Cytometry: SHSY-5Y cells. IHC-P: FFPE human cerebral cortex and human heart muscle tissues.
  • General notes

    This antibody clone is manufactured by Abcam.

    If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

Properties

Applications

Our Abpromise guarantee covers the use of ab77924 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
IHC-Fr Use at an assay dependent concentration. PubMed: 20689587
IHC-P Use a concentration of 1 - 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Flow Cyt Use 1µg for 106 cells.

ab170192 - Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.

WB Use a concentration of 5 µg/ml. Detects a band of approximately 55 kDa (predicted molecular weight: 52 kDa).

Abcam recommends using 1-3% Milk as the blocking agent.  Higher percentage blocking solutions may not give optimal results.

IP Use at an assay dependent concentration.

Target

  • Function
    Functions within a multiprotein E3 ubiquitin ligase complex, catalyzing the covalent attachment of ubiquitin moieties onto substrate proteins, such as BCL2, SYT11, CCNE1, GPR37, STUB1, a 22 kDa O-linked glycosylated isoform of SNCAIP, SEPT5, ZNF746 and AIMP2. Mediates monoubiquitination as well as 'Lys-48'-linked and 'Lys-63'-linked polyubiquitination of substrates depending on the context. Participates in the removal and/or detoxification of abnormally folded or damaged protein by mediating 'Lys-63'-linked polyubiquitination of misfolded proteins such as PARK7: 'Lys-63'-linked polyubiquitinated misfolded proteins are then recognized by HDAC6, leading to their recruitment to aggresomes, followed by degradation. Mediates 'Lys-63'-linked polyubiquitination of SNCAIP, possibly playing a role in Lewy-body formation. Mediates monoubiquitination of BCL2, thereby acting as a positive regulator of autophagy. Promotes the autophagic degradation of dysfunctional depolarized mitochondria. Mediates 'Lys-48'-linked polyubiquitination of ZNF746, followed by degradation of ZNF746 by the proteasome; possibly playing a role in role in regulation of neuron death. Limits the production of reactive oxygen species (ROS). Loss of this ubiquitin ligase activity appears to be the mechanism underlying pathogenesis of PARK2. May protect neurons against alpha synuclein toxicity, proteasomal dysfunction, GPR37 accumulation, and kainate-induced excitotoxicity. May play a role in controlling neurotransmitter trafficking at the presynaptic terminal and in calcium-dependent exocytosis. Regulates cyclin-E during neuronal apoptosis. May represent a tumor suppressor gene.
  • Tissue specificity
    Highly expressed in the brain including the substantia nigra. Expressed in heart, testis and skeletal muscle. Expression is down-regulated or absent in tumor biopsies, and absent in the brain of PARK2 patients. Overexpression protects dopamine neurons from kainate-mediated apoptosis. Found in serum (at protein level).
  • Pathway
    Protein modification; protein ubiquitination.
  • Involvement in disease
    Defects in PARK2 are a cause of Parkinson disease (PARK) [MIM:168600]. A complex neurodegenerative disorder characterized by bradykinesia, resting tremor, muscular rigidity and postural instability. Additional features are characteristic postural abnormalities, dysautonomia, dystonic cramps, and dementia. The pathology of Parkinson disease involves the loss of dopaminergic neurons in the substantia nigra and the presence of Lewy bodies (intraneuronal accumulations of aggregated proteins), in surviving neurons in various areas of the brain. The disease is progressive and usually manifests after the age of 50 years, although early-onset cases (before 50 years) are known. The majority of the cases are sporadic suggesting a multifactorial etiology based on environmental and genetic factors. However, some patients present with a positive family history for the disease. Familial forms of the disease usually begin at earlier ages and are associated with atypical clinical features.
    Defects in PARK2 are the cause of Parkinson disease type 2 (PARK2) [MIM:600116]; also known as early-onset parkinsonism with diurnal fluctuation (EPDF) or autosomal recessive juvenile Parkinson disease (PDJ). A neurodegenerative disorder characterized by bradykinesia, rigidity, postural instability, tremor, and onset usually befor 40. It differs from classic Parkinson disease by early DOPA-induced dyskinesia, diurnal fluctuation of the symptoms, sleep benefit, dystonia and hyper-reflexia. Dementia is absent. Pathologically, patients show loss of dopaminergic neurons in the substantia nigra, similar to that seen in Parkinson disease; however, Lewy bodies (intraneuronal accumulations of aggregated proteins) are absent.
    Note=Defects in PARK2 may be involved in the development and/or progression of ovarian cancer.
  • Sequence similarities
    Belongs to the RBR family. Parkin subfamily.
    Contains 1 IBR-type zinc finger.
    Contains 2 RING-type zinc fingers.
    Contains 1 ubiquitin-like domain.
  • Domain
    The ubiquitin-like domain binds the PSMD4 subunit of 26S proteasomes.
  • Post-translational
    modifications
    Auto-ubiquitinates in an E2-dependent manner leading to its own degradation. Also polyubiquitinated by RNF41 for proteasomal degradation.
    S-nitrosylated. The inhibition of PARK2 ubiquitin E3 ligase activity by S-nitrosylation could contribute to the degenerative process in PD by impairing the ubiquitination of PARK2 substrates.
  • Cellular localization
    Cytoplasm > cytosol. Nucleus. Endoplasmic reticulum. Mitochondrion. Mainly localizes in the cytosol. Co-localizes with SYT11 in neutrites. Co-localizes with SNCAIP in brainstem Lewy bodies. Relocates to dysfunctional mitochondria that have lost the mitochondial membrane potential; recruitement to mitochondria is PINK1-dependent.
  • Information by UniProt
  • Database links
  • Alternative names
    • AR JP antibody
    • E3 ubiquitin ligase antibody
    • E3 ubiquitin protein ligase parkin antibody
    • E3 ubiquitin-protein ligase parkin antibody
    • FRA6E antibody
    • LPRS 2 antibody
    • LPRS2 antibody
    • PARK 2 antibody
    • Park2 antibody
    • Parkin 2 antibody
    • Parkinson disease (autosomal recessive juvenile) 2 antibody
    • Parkinson disease (autosomal recessive, juvenile) 2, parkin antibody
    • Parkinson disease protein 2 antibody
    • Parkinson juvenile disease protein 2 antibody
    • Parkinson protein 2 E3 ubiquitin protein ligase antibody
    • Parkinson protein 2, E3 ubiquitin protein ligase (parkin) antibody
    • PDJ antibody
    • PRKN 2 antibody
    • PRKN antibody
    • PRKN2 antibody
    • PRKN2_HUMAN antibody
    • Ubiquitin E3 ligase PRKN antibody
    see all

Images

  • All lanes : Anti-Parkin antibody [PRK8] (ab77924) at 1/500 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate at 40 µg
    Lane 2 : PARK2 (Parkin) knockout HAP1 whole cell lysate at 40 µg
    Lane 3 : HeLa whole cell lysate at 20 µg
    Lane 4 : Mouse brain whole cell lysate at 20 µg

    Predicted band size: 52 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab77924 observed at 52 kDa. Red - loading control, ab181602, observed at 37 kDa.

    ab77924 was shown to specifically react with Parkin in wild-type HAP1 cells as signal was lost in PARK2 (Parkin) knockout cells. Wild-type and PARK2 (Parkin) knockout samples were subjected to SDS-PAGE. Ab77924 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 1/500 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

    (Parkin expression in HeLa is expected to be negative/low PMID: 14614460 PMID: 12972409)

  • IHC image of Parkin staining in a formalin fixed, paraffin embedded normal human cerebral cortex tissue section, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab77924, 5 µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

     

     

  • IHC image of Parkin staining in a formalin fixed, paraffin embedded normal human heart muscle tissue section*, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab77924, 5 µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

  • Immunohistochemical analysis of transgenic Zebrafish expressing Human Parkin, staining Parkin with ab77924.

    Tissue was fixed with paraformaldehyde and blocked with 10% newborn calf serum with 0.1% Tween for 2 hours. Samples were incubated with primary antibody (1/200) overnight at 4°C. An AlexaFluor®488-conjugated anti-mouse IgG was used as the secondary antibody.
  • All lanes : Anti-Parkin antibody [PRK8] (ab77924) at 1/2000 dilution

    Lane 1 : SHSY-5Y (Human neuroblastoma cell line) Whole Cell Lysate
    Lane 2 : Brain (Rat) Tissue Lysate
    Lane 3 : Brain (Mouse) Tissue Lysate
    Lane 4 : Human brain tissue lysate - total protein (ab29466)

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat polyclonal Secondary Antibody to Mouse IgG - H&L (HRP), pre-adsorbed at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size: 52 kDa
    Additional bands at: 55 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 20 minutes


    All lanes blocked with 3% milk.

  • Overlay histogram showing SH-SY5Y cells stained with ab77924 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab77924, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in SH-SY5Y cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

References

This product has been referenced in:
  • Gu X  et al. Lead (Pb) induced ATM-dependent mitophagy via PINK1/Parkin pathway. Toxicol Lett 291:92-100 (2018). ICC/IF . Read more (PubMed: 29660402) »
  • da Silva-Camargo CCV  et al. Parkin protein expression and its impact on survival of patients with advanced colorectal cancer. Cancer Biol Med 15:61-69 (2018). IHC-P ; Human . Read more (PubMed: 29545969) »
See all 25 Publications for this product

Customer reviews and Q&As

1-10 of 15 Abreviews or Q&A

Answer

Thank you for your response.
As our Abpromise indicates, in the event that a product is not functioning in the applications/species cited on the product data sheet (and the problem has been reported within 6 months of purchase) we will happily offer a credit note/refund to the value of the product purchased.
These two antibodies (ab15954 and ab77924) have been tested in mouse samples but no cross-reactivity data available for canine yet.
ab15954: tested in Mouse, Rat, Cow, Human, Monkey
positive control: HEK293, SH-SY5Y or COS7 cell lysate
ab77924 : tested in Mouse, Rat, Human;
positive control: HEK293 cell lysate. Mouse brain tissue.
There are differences in the expressed levels of Parkin in different tissues and cells so I would advise you to consult this site; you may get some useful information: http://www.proteinatlas.org/ENSG00000185345
During characterization, cell lysate from HEK293 cells was used so I would advise you to run this type of lysate or any recommended lysates listed on the product datasheets as positive control alongside with the samples.
I hope this will be useful for you.
Should you still have any problem with this antibody after following these suggestions, then please do not hesitate to contact our Technical Department again (confirming the date of purchase, Abcam Order Number and the address of your institute).

Read More
Application
Western blot
Sample
Human Cell lysate - other (head and neck cancer)
Gel Running Conditions
Reduced Denaturing (12% SDS page gel)
Loading amount
30 µg
Treatment
5uM DIM-C-pPhtBu for 24h
Specification
head and neck cancer
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Dr. Sung Un Kang

Verified customer

Submitted Sep 18 2018

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Tissue lysate - whole (liver)
Gel Running Conditions
Reduced Denaturing
Loading amount
25 µg
Specification
liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Aug 02 2016

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
Sample
Chinese Hamster Cell (CHO cell)
Specification
CHO cell
Permeabilization
Yes - 1% TRITON-X-100
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Mar 18 2015

Application
Western blot
Loading amount
25 µg
Gel Running Conditions
Reduced Denaturing
Sample
Mouse Cell lysate - whole cell (liver, hepatocytes)
Specification
liver, hepatocytes
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jan 07 2015

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - other (brain mitochondria)
Loading amount
40 µg
Specification
brain mitochondria
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Feb 11 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Drosophila melanogaster Tissue lysate - whole (fly heart tube)
Gel Running Conditions
Reduced Denaturing
Loading amount
10 µg
Specification
fly heart tube
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: rt°C

Abcam user community

Verified customer

Submitted Feb 11 2013

Application
Immunocytochemistry/ Immunofluorescence
Sample
Drosophila melanogaster Cell (fly heart tube)
Permeabilization
Yes - 1% triton x100
Specification
fly heart tube
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: rt°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Feb 08 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunoprecipitation
Sample
Mouse Tissue lysate - whole (mouse heart)
Total protein in input
200 µg
Specification
mouse heart
Immuno-precipitation step
Protein G

Abcam user community

Verified customer

Submitted Feb 08 2013

Answer

Vielen Dank nochmal für Ihre Anfrage.



Leider bieten wir generell keine kostenlosen Proben zu Testzwecken an, da wir bei über 100.000Produkten gar nicht die logistischen Möglichkeiten haben, neben unserer eigentlichen Produktpallette Teströhrchen zu lagern. Hier bei Abcam verfahren wir so, dass wir einen kostenlosen Ersatz oder eine Gutschrift verschicken, falls ein Produkt nicht so funktioniert wie auf unserem Datenblatt beschrieben, und er innerhalb der 180Tage Garantiezeit reklamiert wird.
Wir bieten auch generell keine Institutsrabatte, aber wir möchten Sie jedoch einladen, unser Abreview-System zu verwenden.


Falls sich jetzt herausstellt, dass die Tipps, die ich Ihnen in der vorangegangenen Email gegeben habe habe nicht funktionieren, bzw der AK im Chemolumineszenz Experiment die gleichen Ergebnisse zeigt, werden wir Ihnen gerne einen kostenlosen Ersatz zusenden, gerne auch einen alternativen Antikörper.



Ich hoffe, dies hilft Ihnen weiter. Bitte zögern Sie nicht, sich wieder bei uns zu melden, falls Sie weitere Fragen haben.

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1-10 of 15 Abreviews or Q&A

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