Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

Reviews (3) Submit a question References (10)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR18351] to SQSTM1 / p62
  • Suitable for: Flow Cyt, IHC-P, WB, ICC/IF, IP
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-SQSTM1 / p62 antibody [EPR18351]
    See all SQSTM1 / p62 primary antibodies

  • Description

    Rabbit monoclonal [EPR18351] to SQSTM1 / p62

  • Host species

    Rabbit

  • Tested applications

    Suitable for: Flow Cyt, IHC-P, WB, ICC/IF, IPmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant full length protein within Human SQSTM1/ p62 aa 1 to the C-terminus. The exact sequence is proprietary.
    Database link: Q13501

  • Positive control

    • WB: Human liver and kidney lysates; HepG2 and HeLa whole cell lysates. IHC-P: Human hepatocellular carcinoma and lung cancer tissues. ICC/IF: HeLa cells (untreated and treated with chloroquine), HAP1 cells (untreated and treated with chloroquine) Flow Cyt: PC-3 cells. IP: HeLa and HepG2 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.

Properties

Applications

Our Abpromise guarantee covers the use of ab207305 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
Flow Cyt 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 
IHC-P 1/2000. 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 62 kDa (predicted molecular weight: 47 kDa).
ICC/IF Use a concentration of 1 µg/ml.
IP 1/40.

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.
  • Information by UniProt

  • Database links

    • 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 [EPR18351] (ab207305)
      Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: SQSTM1/p62 knockout HAP1 cell lysate (20 µg)
      Lane 3: HeLa cell lysate (20 µg)
      Lane 4: HepG2 cell lysate (20 µg)
      Lanes 1 - 4: Merged signal (red and green).

      Green - target observed at 55 kDa. Red - loading control, ab8245, observed at 37 kDa.

      This western blot image is a comparison between ab207305 and a competitor's discontinued goat polyclonal antibody.

       

       

    • Immunocytochemistry/ Immunofluorescence - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunocytochemistry/ Immunofluorescence - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      ab207305 staining SQSTM1 in wild-type HAP1 cells and knockout cells, untreated and chloroquine-treated (50μM, 24 hours). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab207305 at 1μg/ml and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

      Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

    • Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: SQSTM1/p62 knockout HAP1 cell lysate (20 µg)
      Lane 3: HeLa cell lysate (20 µg)
      Lane 4: HepG2 cell lysate (20 µg)
      Lanes 1 - 4: Merged signal (red and green). Green - ab207305 observed at 55 kDa. Red - loading control, ab8245, observed at 37 kDa.

      ab207305 was shown to specifically react with SQSTM1/p62 in wild-type HAP1 cells. No band was observed when SQSTM1/p62 knockout samples were used. Wild-type and SQSTM1/p62 knockout samples were subjected to SDS-PAGE, ab207305 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma tissue labeling SQSTM1 / p62 with ab207305 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on human hepatocellular carcinoma is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunocytochemistry/ Immunofluorescence - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunocytochemistry/ Immunofluorescence - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      ab207305 staining SQSTM1/p62 in HeLa cells +/- Chloroquine (50μM, 24 hours). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab207305 at 1μg/ml and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 1/250 dilution (shown in pseudocolor red) followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

      Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

    • Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      All lanes : Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305) at 1/1000 dilution

      Lane 1 : Human liver lysate
      Lane 2 : Human kidney lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

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


      Exposure time: 30 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

      The MW observed is consistent with what has been described in the literature (PMID: PMC4344198).

    • Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Western blot - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      All lanes : Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305) at 1/5000 dilution

      Lane 1 : HepG2 (Human liver hepatocellular carcinoma) whole cell lysate
      Lane 2 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell 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: 47 kDa
      Observed band size: 62 kDa why is the actual band size different from the predicted?


      Exposure time: 30 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

      The MW observed is consistent with what has been described in the literature (PMID: PMC4344198).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Immunohistochemical analysis of paraffin-embedded human lung cancer tissue labeling SQSTM1 / p62 with ab207305 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm and weakly nucleus staining on human lung cancer is observed. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

      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-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Immunohistochemical analysis of paraffin-embedded human liver tissue labeling SQSTM1 / p62 with ab207305 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Hhman normal liver tissue. Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Flow Cytometry - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Flow Cytometry - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      Flow cytometric analysis of 4% paraformaldehyde-fixed PC-3 (Human prostate adenocarcinoma cell line) cells labeling SQSTM1 / p62 with ab207305 at 1/100 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat Anti-Rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.

    • Immunoprecipitation - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunoprecipitation - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      SQSTM1 / p62 was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab207305 at 1/40 dilution.
      Western blot was performed from the immunoprecipitate using ab207305 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
      Lane 1: HeLa whole cell lysate 10ug (Input).
      Lane 2: ab207305 IP in HeLa whole cell lysate.
      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab207305 in HeLa whole cell lysate.
      Blocking and dilution buffer and concentration: 5% NFDM/TBST.
      Exposure time: 1 second.

    • Immunoprecipitation - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)
      Immunoprecipitation - Anti-SQSTM1 / p62 antibody [EPR18351] (ab207305)

      SQSTM1 / p62 was immunoprecipitated from 1mg of HepG2 (Human liver hepatocellular carcinoma) whole cell lysate with ab207305 at 1/40 dilution.
      Western blot was performed from the immunoprecipitate using ab207305 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
      Lane 1: HepG2 whole cell lysate 10ug (Input).
      Lane 2: ab207305 IP in HepG2 whole cell lysate.
      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab207305 in HepG2 whole cell lysate.
      Blocking and dilution buffer and concentration: 5% NFDM/TBST.
      Exposure time: 1 second.

    References

    Publishing research using ab207305? Please let us know so that we can cite the reference in this datasheet.

    ab207305 has been referenced in 10 publications.

    • Zhang J  et al. p62 functions as an oncogene in colorectal cancer through inhibiting apoptosis and promoting cell proliferation by interacting with the vitamin D receptor. Cell Prolif N/A:e12585 (2019). PubMed: 30793399
    • Lin P  et al. An autophagy-related gene expression signature for survival prediction in multiple cohorts of hepatocellular carcinoma patients. Oncotarget 9:17368-17395 (2018). PubMed: 29707114
    • Chen C  et al. LncRNA HULC mediates radioresistance via autophagy in prostate cancer cells. Braz J Med Biol Res 51:e7080 (2018). PubMed: 29694502
    • Xu J  et al. Overexpression of long non-coding RNA H19 promotes invasion and autophagy via the PI3K/AKT/mTOR pathways in trophoblast cells. Biomed Pharmacother 101:691-697 (2018). WB . PubMed: 29522949
    • Liang L  et al. Investigation of cancer-associated fibroblasts and p62 expression in oral cancer before and after chemotherapy. J Craniomaxillofac Surg 46:605-610 (2018). IHC-P . PubMed: 29439841
    • Zhu J  et al. Beclin1 overexpression suppresses tumor cell proliferation and survival via an autophagy-dependent pathway in human synovial sarcoma cells. Oncol Rep 40:1927-1936 (2018). PubMed: 30066884
    • Xiao T  et al. RACK1 promotes tumorigenicity of colon cancer by inducing cell autophagy. Cell Death Dis 9:1148 (2018). PubMed: 30451832
    • Zhu JF  et al. Annexin A1-suppressed autophagy promotes nasopharyngeal carcinoma cell invasion and metastasis by PI3K/AKT signaling activation. Cell Death Dis 9:1154 (2018). PubMed: 30459351
    • Li Y  et al. Octreotide Alleviates Autophagy by Up-Regulation of MicroRNA-101 in Intestinal Epithelial Cell Line Caco-2. Cell Physiol Biochem 49:1352-1363 (2018). PubMed: 30205395
    • Lei W  et al. Chlamydia trachomatis plasmid-encoded protein pORF5 protects mitochondrial function by inducing mitophagy and increasing HMGB1 expression. Pathog Dis 75:N/A (2017). PubMed: 29040500

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-3 of 3 Abreviews or Q&A

    Immunoprecipitation abreview for Anti-SQSTM1 / p62 antibody [EPR18351]

     Excellent
    Abreviews
    abreview image
    Application
    Immunoprecipitation
    Sample
    Human Cell lysate - whole cell (human kidney)
    Total protein in input
    200 µg
    Immuno-precipitation step
    Protein A
    Specification
    human kidney
    Read More

    Abcam user community

    Verified customer

    Submitted Sep 06 2018

    Immunoprecipitation abreview for Anti-SQSTM1 / p62 antibody [EPR18351]

     Excellent
    Abreviews
    abreview image
    Application
    Immunoprecipitation
    Sample
    Mouse Cell lysate - whole cell (Mouse Embyronic fibroblasts)
    Total protein in input
    200 µg
    Immuno-precipitation step
    Protein A
    Specification
    Mouse Embyronic fibroblasts
    Read More

    Abcam user community

    Verified customer

    Submitted Aug 22 2017

    Immunocytochemistry/ Immunofluorescence abreview for Anti-SQSTM1 / p62 antibody [EPR18351]

     Excellent
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Mouse Cell (Mouse Embyronic fibroblasts)
    Permeabilization
    Yes - Digitonin
    Specification
    Mouse Embyronic fibroblasts
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 21°C
    Fixative
    Formaldehyde
    Read More

    Dr. Ryan Russell

    Verified customer

    Submitted Aug 10 2017

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com