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  1. Link

    sqstm1--p62-antibody-3p62-lck-ligand-ab280086.pdf

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Validated using a knockout cell lineRecombinant

Recombinant Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

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Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
  • Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
  • Immunocytochemistry - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
  • Immunoprecipitation - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Mouse monoclonal [3/P62 LCK LIGAND] to SQSTM1 / p62
  • Suitable for: ICC, IP, WB, IHC-P
  • Knockout validated
  • Reacts with: Human

Conjugates logo Related conjugates and formulations

Carrier Free

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Overview

  • Product name

    Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND]
    See all SQSTM1 / p62 primary antibodies
  • Description

    Mouse monoclonal [3/P62 LCK LIGAND] to SQSTM1 / p62
  • Host species

    Mouse
  • Tested applications

    Suitable for: ICC, IP, WB, IHC-Pmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: Wild-type HAP1; HeLa; MCF7; HEK-293 whole cell lysates. IHC-P: Human stomach carcinoma, lung carcinoma tissue. ICC: HeLa cells. IP: HeLa 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.

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: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    3/P62 LCK LIGAND
  • Isotype

    IgG1
  • Light chain type

    kappa
  • Research areas

    • Signal Transduction
    • Signaling Pathway
    • Nuclear Signaling
    • NFkB Pathway
    • Signal Transduction
    • Protein Trafficking
    • Vesicle Transport
    • Regulation
    • Epigenetics and Nuclear Signaling
    • Transcription
    • Polymerase associated factors
    • Pol II Transcription
    • Other
    • Cardiovascular
    • Heart
    • Autophagy
    • Autophagosome
    • Metabolism
    • Pathways and Processes
    • Metabolism processes
    • Autophagy and mitophagy
    • Autophagosome
    • Cancer
    • Cell Death
    • Autophagy
    • Autophagosome
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] - BSA and Azide free (ab280091)
  • Compatible Secondaries

    • Rabbit monoclonal [M1gG51-4] Anti-Mouse IgG1 H&L (ab125913)
    • VeriBlot for IP Detection Reagent (HRP) (ab131366)
    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
  • Dyes/Markers

    • Anti-beta Tubulin antibody [EPR16774] (ab179513)
  • Isotype control

    • Mouse IgG1, kappa monoclonal [MOPC-21] - isotype control (ab18443)
  • Related Products

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) (ab150080)
    • Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab280086 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
ICC
1/50.
IP
1/30.
WB
1/1000. Predicted molecular weight: 47 kDa.
IHC-P
1/1000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Notes
ICC
1/50.
IP
1/30.
WB
1/1000. Predicted molecular weight: 47 kDa.
IHC-P
1/1000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Target

  • Function

    Adapter protein which binds ubiquitin and may regulate the activation of NFKB1 by TNF-alpha, nerve growth factor (NGF) and interleukin-1. May play a role in titin/TTN downstream signaling in muscle cells. May regulate signaling cascades through ubiquitination. Adapter that mediates the interaction between TRAF6 and CYLD (By similarity). May be involved in cell differentiation, apoptosis, immune response and regulation of K(+) channels.
  • Tissue specificity

    Ubiquitously expressed.
  • Involvement in disease

    Defects in SQSTM1 are a cause of Paget disease of bone (PDB) [MIM:602080]. PDB is a metabolic bone disease affecting the axial skeleton and characterized by focal areas of increased and disorganized bone turn-over due to activated osteoclasts. Manifestations of the disease include bone pain, deformity, pathological fractures, deafness, neurological complications and increased risk of osteosarcoma. PDB is a chronic disease affecting 2 to 3% of the population above the age of 40 years.
  • Sequence similarities

    Contains 1 OPR domain.
    Contains 1 UBA domain.
    Contains 1 ZZ-type zinc finger.
  • Domain

    The UBA domain binds specifically 'Lys-63'-linked polyubiquitin chains of polyubiquitinated substrates. Mediates the interaction with TRIM55.
    The OPR domain mediates homooligomerization and interactions with PRKCZ, PRKCI, MAP2K5 and NBR1.
    The ZZ-type zinc finger mediates the interaction with RIPK1.
  • Post-translational
    modifications

    Phosphorylated. May be phosphorylated by PRKCZ (By similarity). Phosphorylated in vitro by TTN.
  • Cellular localization

    Cytoplasm. Late endosome. Nucleus. Sarcomere (By similarity). In cardiac muscles localizes to the sarcomeric band (By similarity). Localizes to late endosomes. May also localize to the nucleus. Accumulates in neurofibrillary tangles and in Lewy bodies of neurons from individuals with Alzheimer and Parkinson disease respectively. Enriched in Rosenthal fibers of pilocytic astrocytoma. In liver cells, accumulates in Mallory bodies associated with alcoholic hepatitis, Wilson disease, indian childhood cirrhosis and in hyaline bodies associated with hepatocellular carcinoma.
  • Target information above from: UniProt accession Q13501 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 8878 Human
    • Omim: 601530 Human
    • SwissProt: Q13501 Human
    • Unigene: 709030 Human
    • Alternative names

      • A170 antibody
      • DMRV antibody
      • EBI 3 associated protein of 60 kDa antibody
      • EBI 3 associated protein p60 antibody
      • EBI3 associated protein of 60 kDa antibody
      • EBI3 associated protein p60 antibody
      • EBI3-associated protein of 60 kDa antibody
      • EBIAP antibody
      • FTDALS3 antibody
      • MGC127197 antibody
      • ORCA antibody
      • OSF-6 antibody
      • Osi antibody
      • OSIL antibody
      • Oxidative stress induced like antibody
      • p60 antibody
      • p62 antibody
      • p62B antibody
      • Paget disease of bone 3 antibody
      • PDB 3 antibody
      • PDB3 antibody
      • Phosphotyrosine independent ligand for the Lck SH2 domain of 62 kDa antibody
      • Phosphotyrosine independent ligand for the Lck SH2 domain p62 antibody
      • Phosphotyrosine-independent ligand for the Lck SH2 domain of 62 kDa antibody
      • PKC-zeta-interacting protein antibody
      • Protein kinase C-zeta-interacting protein antibody
      • Sequestosome 1 antibody
      • Sequestosome-1 antibody
      • SQSTM 1 antibody
      • SQSTM_HUMAN antibody
      • Sqstm1 antibody
      • STAP antibody
      • STONE14 antibody
      • Ubiquitin binding protein p62 antibody
      • Ubiquitin-binding protein p62 antibody
      • ZIP 3 antibody
      • ZIP antibody
      • ZIP3 antibody
      see all

    Images

    • Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      All lanes : Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086) at 1/1000 dilution

      Lane 1 : Wild-type HAP1 (human chronic myelogenous leukemia near-haploid cell line), whole cell lysate
      Lane 2 : SQSTM1 knockout HAP1 (human chronic myelogenous leukemia near-haploid cell line), whole cell lysate
      Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate
      Lane 4 : MCF7 (human breast adenocarcinoma epithelial cell), whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Mouse IgG H&L (IRDye® 800CW) (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) (ab216777) at 1/10000 dilution

      Predicted band size: 47 kDa
      Observed band size: 62 kDa why is the actual band size different from the predicted?



      Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS

      Lanes 1 - 4: Merged signal (red and green). Green - ab280086 observed at 62 kDa. Red - loading control ab181602 (Rabbit monoclonal [EPR16891] to GAPDH) observed at 36 kDa.

      Lanes 1-2: ab280086 Anti-SQSTM1/p62 antibody was shown to react with SQSTM1 in HAP1 cells in Western blot. Loss of signal was observed when SQSTM1 knockout sample was used. Wild-type and SQSTM1 knockout samples were subjected to SDS-PAGE. ab280086 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) were incubated at 4°C overnight at 1/1000 dilution and 1/20000 dilution respectively.

      Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680CW) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800RD) preadsorbed (ab216772) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging. 

    • Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Western blot - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      All lanes : Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086) at 1/1000 dilution

      Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell ), whole cell lysate
      Lane 2 : HEK-293 (human embryonic kidney epithelial cell), whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Peroxidase-Conjugated Goat anti-Mouse IgG (H+L) at 1/10000 dilution

      Predicted band size: 47 kDa
      Observed band size: 62 kDa why is the actual band size different from the predicted?



      Blocking and diluting buffer and concentration: 5% NFDM/TBST

      The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 24086455).

      Lysates were made freshly and used in WB immediately to minimize protein degradation.

      Exposure time: 15 seconds

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

      Immunohistochemical analysis of paraffin-embedded Human stomach carcinoma tissue labeling SQSTM1 / p62 with ab280086 at 1/1000 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on human stomach carcinoma. The section was incubated with ab280086 for 30 mins at room temperature and followed by mouse specific IgG antibody (ab125913) for 8mins. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

      Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.

      Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

      Immunohistochemical analysis of paraffin-embedded Human lung carcinoma tissue labeling SQSTM1 / p62 with ab280086 at 1/1000 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on human lung carcinoma. The section was incubated with ab280086 for 30 mins at room temperature and followed by mouse specific IgG antibody (ab125913) for 8mins. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

      Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.

      Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

    • Immunocytochemistry - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Immunocytochemistry - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

      Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa cells labelling SQSTM1 / p62 with ab280086 at 1/50 dilution, followed by ab150113 Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HeLa cell line. ab179513 Anti-beta Tubulin rabbit monoclonal antibody was used to counterstain tubulin at 1/200 dilution followed by ab150080 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594)(Red). The Nuclear counterstain was DAPI (Blue).

      Secondary antibody only control: Secondary antibody is ab150113 Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.

    • Immunoprecipitation - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)
      Immunoprecipitation - Anti-SQSTM1 / p62 antibody [3/P62 LCK LIGAND] (ab280086)

      SQSTM1 / p62 was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell ), whole cell lysate 10 ug with ab280086 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab280086 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

      Lane 1: HeLa (human cervix adenocarcinoma epithelial cell ), whole cell lysate 10 ug

      Lane 2: ab280086 IP in HeLa whole cell lysate

      Lane 3: Mouse monoclonal IgG1(ab18443) instead of ab280086 in HeLa whole cell lysate

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

      Exposure time: 26 seconds

       

    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.

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (0)

    Publishing research using ab280086? Please let us know so that we can cite the reference in this datasheet.

    ab280086 has not yet been referenced specifically in any publications.

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