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  1. Link

    shp2-antibody-y478-ab32083.pdf

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Signal Transduction Protein Phosphorylation Tyrosine Phosphatases
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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-SHP2 antibody [Y478] (ab32083)

  • Datasheet
  • SDS
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Western blot - Anti-SHP2 antibody [Y478] (ab32083)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)
  • Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)
  • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
  • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
  • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
  • Immunoprecipitation - Anti-SHP2 antibody [Y478] (ab32083)
  • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
  • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)
  • Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)
  • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
  • Anti-SHP2 antibody [Y478] (ab32083)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y478] to SHP2
  • Suitable for: WB, IHC-P, IP, Flow Cyt (Intra), ICC/IF
  • Knockout validated
  • Reacts with: Human

Conjugates logo Related conjugates and formulations

Alexa Fluor® 594 APC Carrier Free HRP PE

You may also be interested in

Reagent
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VeriBlot for IP Detection Reagent (HRP) (ab131366)
Secondary
Product image
Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
Knockout
Product image
Human PTPN11 (SHP2) knockout HEK-293T cell line (ab266450)

View more associated products

Overview

  • Product name

    Anti-SHP2 antibody [Y478]
    See all SHP2 primary antibodies
  • Description

    Rabbit monoclonal [Y478] to SHP2
  • Host species

    Rabbit
  • Specificity

    ab32083 recognises SHP2. This antibody is predicted to detect splice isoform 2 based on sequence analysis.
  • Tested applications

    Suitable for: WB, IHC-P, IP, Flow Cyt (Intra), ICC/IFmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human SHP2 aa 500-600 (C terminal). The exact sequence is proprietary.

  • Positive control

    • IHC-P: Human breast carcinoma and endometrium tissue. WB: HEK-293T, Jurkat, and THP-1 cell lysate. ICC/IF: Hek293 and A431 cells. Flow Cyt (intra): HAP1-WT and Jurkat cells IP: THP-1 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.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

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: 40% Glycerol (glycerin, glycerine), 59% PBS, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    Y478
  • Isotype

    IgG
  • Research areas

    • Signal Transduction
    • Protein Phosphorylation
    • Tyrosine Phosphatases
    • Neuroscience
    • Sensory System
    • Auditory system

Associated products

  • Alternative Versions

    • Anti-SHP2 antibody [Y478] - BSA and Azide free (ab182179)
    • Alexa Fluor® 594 Anti-SHP2 antibody [Y478] (ab210616)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081)
    • Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Conjugation kits

    • PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • KO cell lines

    • Human PTPN11 (SHP2) knockout HEK-293T cell line (ab266450)
  • KO cell lysates

    • Human PTPN11 (SHP2) knockout HEK-293T cell lysate (ab257618)
  • KO cell pellets

    • Human PTPN11 (SHP2) knockout HEK-293T cell pellet (ab278893)
  • Positive Controls

    • Jurkat whole cell lysate (ab7899)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab32083 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
WB
1/1000 - 1/10000. Predicted molecular weight: 68 kDa.
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IP
1/40.

For unpurified use at 1/50

Flow Cyt (Intra)
1/50.

For unpurified use at 0.1 µg/ml

ICC/IF
1/50.

For unpurified use at 1/100.

Notes
WB
1/1000 - 1/10000. Predicted molecular weight: 68 kDa.
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IP
1/40.

For unpurified use at 1/50

Flow Cyt (Intra)
1/50.

For unpurified use at 0.1 µg/ml

ICC/IF
1/50.

For unpurified use at 1/100.

Target

  • Function

    Acts downstream of various receptor and cytoplasmic protein tyrosine kinases to participate in the signal transduction from the cell surface to the nucleus.
  • Tissue specificity

    Widely expressed, with highest levels in heart, brain, and skeletal muscle.
  • Involvement in disease

    Defects in PTPN11 are the cause of LEOPARD syndrome type 1 (LEOPARD1) [MIM:151100]. It is an autosomal dominant disorder allelic with Noonan syndrome. The acronym LEOPARD stands for lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonic stenosis, abnormalities of genitalia, retardation of growth, and deafness.
    Defects in PTPN11 are the cause of Noonan syndrome type 1 (NS1) [MIM:163950]. Noonan syndrome (NS) is a disorder characterized by dysmorphic facial features, short stature, hypertelorism, cardiac anomalies, deafness, motor delay, and a bleeding diathesis. Some patients with Noonan syndrome type 1 develop multiple giant cell lesions of the jaw or other bony or soft tissues, which are classified as pigmented villomoduolar synovitis (PVNS) when occurring in the jaw or joints. Note=Mutations in PTPN11 account for more than 50% of the cases. Rarely, NS is associated with juvenile myelomonocytic leukemia (JMML). NS1 inheritance is autosomal dominant.
    Defects in PTPN11 are a cause of juvenile myelomonocytic leukemia (JMML) [MIM:607785]. JMML is a pediatric myelodysplastic syndrome that constitutes approximately 30% of childhood cases of myelodysplastic syndrome (MDS) and 2% of leukemia. It is characterized by leukocytosis with tissue infiltration and in vitro hypersensitivity of myeloid progenitors to granulocyte-macrophage colony stimulating factor.
    Defects in PTPN11 are a cause of metachondromatosis (MC) [MIM:156250]. It is a skeletal disorder with radiologic fetarures of both multiple exostoses and Ollier disease, characterized by the presence of multiple enchondromas and osteochondroma-like lesions.
  • Sequence similarities

    Belongs to the protein-tyrosine phosphatase family. Non-receptor class 2 subfamily.
    Contains 2 SH2 domains.
    Contains 1 tyrosine-protein phosphatase domain.
  • Domain

    The SH2 domains repress phosphatase activity. Binding of these domains to phosphotyrosine-containing proteins relieves this auto-inhibition, possibly by inducing a conformational change in the enzyme.
  • Post-translational
    modifications

    Phosphorylated on Tyr-546 and Tyr-584 upon receptor protein tyrosine kinase activation; which creates a binding site for GRB2 and other SH2-containing proteins.
  • Cellular localization

    Cytoplasm.
  • Target information above from: UniProt accession Q06124 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 5781 Human
    • Omim: 176876 Human
    • SwissProt: Q06124 Human
    • Unigene: 506852 Human
    • Alternative names

      • BPTP3 antibody
      • CFC antibody
      • JMML antibody
      • METCDS antibody
      • MGC14433 antibody
      • NS1 antibody
      • OTTHUMP00000166107 antibody
      • OTTHUMP00000166108 antibody
      • Protein tyrosine phosphatase 2 antibody
      • Protein tyrosine phosphatase 2C antibody
      • Protein tyrosine phosphatase non receptor type 11 antibody
      • Protein-tyrosine phosphatase 1D antibody
      • Protein-tyrosine phosphatase 2C antibody
      • PTN11_HUMAN antibody
      • PTP-1D antibody
      • PTP-2C antibody
      • PTP1D antibody
      • PTP2C antibody
      • PTPN11 antibody
      • SAP2 antibody
      • SH-PTP2 antibody
      • SH-PTP3 antibody
      • SH2 domain containing protein tyrosine phosphatase 2 antibody
      • SHP 2 antibody
      • SHP-2 antibody
      • Shp2 antibody
      • SHPTP2 antibody
      • SHPTP3 antibody
      • Syp antibody
      • Tyrosine-protein phosphatase non-receptor type 11 antibody
      see all

    Images

    • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      All lanes : Anti-SHP2 antibody [Y478] (ab32083) at 1/1000 dilution

      Lane 1 : Wild-type HEK-293T cell lysate
      Lane 2 : PTPN11 knockout HEK-293T cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 68 kDa
      Observed band size: 68 kDa



      Lanes 1- 2: Merged signal (red and green). Green - ab32083 observed at 68 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

       ab32083 was shown to react with SHP2 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266450 (knockout cell lysate ab257618) was used. Wild-type HEK-293T and PTPN11 knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32083 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human endometrium cancer tissue sections labeling SHP2 with Purified ab32083 at 1:100 dilution (5.51 µg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0)ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody.Negative control:PBS instead of the primary antibody.Hematoxylinwas used as a counterstain
    • Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)
      Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)
      Immunocytochemistry/ Immunofluorescence analysis of A431 (Human epidermoid carcinoma epithelial cell) cells labeling SHP2 with Purified ab32083 at 1:50 dilution (11 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor®594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor®488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
    • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
      Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)

      Intracellular Flow Cytometry analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling SHP2 with purified ab32083 at 1/50 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

       

    • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Western blot - Anti-SHP2 antibody [Y478] (ab32083)

      Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: SHP2 knockout HAP1 cell lysate (20 µg)
      Lane 3: A431 cell lysate (20 µg)
      Lane 4: Jurkat cell lysate (20 µg)
      Lanes 1 to 4: Merged signal (red and green). Green - ab32083 observed at 68 kDa. Red - loading control, ab8245, observed at 37 kDa.
      unpurified ab32083 was shown to specifically react with SHP2 when SHP2 knockout samples were used. Wild-type and SHP2 knockout samples were subjected to SDS-PAGE. ab32083 and ab8245 (loading control to GAPDH) were both 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 h at room temperature before imaging.

    • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Anti-SHP2 antibody [Y478] (ab32083) at 0.3 µg/ml (purified) + THP-1 (Human monocytic leukemia monocyte) whole cell lysates at 15 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 68 kDa



      Blocking and diluting buffer: 5% NFDM/TBST. 

    • Immunoprecipitation - Anti-SHP2 antibody [Y478] (ab32083)
      Immunoprecipitation - Anti-SHP2 antibody [Y478] (ab32083)

      ab32083 (purified) at 1:40 dilution (2µg) immunoprecipitating SHP2 in THP-1 whole cell lysate.
      Lane 1 (input): THP-1 (Human monocytic leukemia monocyte) whole cell lysate 10µg
      Lane 2 (+): ab32083 & THP-1 whole cell lysate
      Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32083 in THP-1 whole cell lysate
      For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
      Blocking and diluting buffer: 5% NFDM/TBST.

    • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
      Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)

      Overlay histogram showing HAP1 wildtype (green line) and HAP1-PTPN11 knockout cells (red line) stained with ab32083. The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (unpurified ab32083, 0.1µg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) presorbed (ab150081) at 1/2000 dilution for 30 min at 22°C. A rabbit IgG isotype control antibody (ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-PTPN11 knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity). Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter. This antibody can also be used in HAP1 cells fixed with 80% methanol (5 min) permeabilized with 0.1% PBS-Triton X-100 for 15 min under the same conditions. 

       

       

    • Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)
      Flow Cytometry (Intracellular) - Anti-SHP2 antibody [Y478] (ab32083)

      Intracellular Flow Cytometry analysis of Jurkat (human acute T cell leukemia) cells labeling SHP2 with unpurified ab32083 at 1/500 dilution (1ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488) (ab150077) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) (ab172730) was used as the isotype control, Cell without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control. 

       

       

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SHP2 antibody [Y478] (ab32083)

      Immunohistochemical analysis of SHP2 expression in paraffin embedded human breast carcinoma, using 1/50 unpurified ab32083.

    • Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)
      Immunocytochemistry/ Immunofluorescence - Anti-SHP2 antibody [Y478] (ab32083)

      unpurified ab32083 stained Hek293 cells. The cells were 100% methanol fixed for 5 minutes at -20°C and then incubated in 1%BSA / 10% normal goat  serum / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32083 at 1/100 dilution) overnight at +4°C. The secondary antibody (pseudo-colored green) was Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) used at a 1/1000 dilution for 1hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

    • Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Western blot - Anti-SHP2 antibody [Y478] (ab32083)
      Anti-SHP2 antibody [Y478] (ab32083) at 1/5000 dilution (unpurified) + Jurkat cell lysate

      Predicted band size: 68 kDa
      Observed band size: 70 kDa why is the actual band size different from the predicted?

    • Anti-SHP2 antibody [Y478] (ab32083)
      Anti-SHP2 antibody [Y478] (ab32083)

    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 (10)

    Publishing research using ab32083? Please let us know so that we can cite the reference in this datasheet.

    ab32083 has been referenced in 10 publications.

    • Yang H  et al. The clinicopathological and prognostic implications of tyrosine phosphatase SHP2 and ankyrin Hook1 gene expression in non- small cell lung cancer patients treated with gemcitabine plus platinum as first-line chemotherapy. Ann Palliat Med 9:2943-2952 (2020). PubMed: 32819122
    • Tang Y  et al. Bioinformatic analysis of differentially expressed genes and identification of key genes in EBV-transformed lymphoblasts. Biomed Pharmacother 116:108984 (2019). PubMed: 31129512
    • Yao D  et al. CD47 is associated with the up-regulation of the PD-1 oncogenic signaling pathway. Int J Clin Exp Pathol 11:5612-5621 (2018). PubMed: 31949648
    • Zhang T  et al. Toosendanin demonstrates promising antitumor efficacy in osteosarcoma by targeting STAT3. Oncogene 36:6627-6639 (2017). PubMed: 28783167
    • Peng YC  et al. Combination of 5-fluorouracil and 2-morphilino-8-phenyl-4H-chromen-4-one may inhibit liver cancer stem cell activity. Tumour Biol N/A:N/A (2016). Human . PubMed: 26886287
    • Maeshima K  et al. Abnormal PTPN11 enhancer methylation promotes rheumatoid arthritis fibroblast-like synoviocyte aggressiveness and joint inflammation. JCI Insight 1:N/A (2016). IHC ; Human . PubMed: 27275015
    • Li K  et al. Genome-Wide Transcriptional Analysis Reveals the Protection against Hypoxia-Induced Oxidative Injury in the Intestine of Tibetans via the Inhibition of GRB2/EGFR/PTPN11 Pathways. Oxid Med Cell Longev 2016:6967396 (2016). WB ; Human . PubMed: 27594973
    • Zheng J  et al. Pancreatic cancer risk variant in LINC00673 creates a miR-1231 binding site and interferes with PTPN11 degradation. Nat Genet 48:747-57 (2016). PubMed: 27213290
    • Zheng J  et al. Expression and prognosis value of SHP2 in patients with pancreatic ductal adenocarcinoma. Tumour Biol N/A:N/A (2015). IHC ; Human . PubMed: 26695153
    • Xie H  et al. Upregulation of Src homology phosphotyrosyl phosphatase 2 (Shp2) expression in oral cancer and knockdown of Shp2 expression inhibit tumor cell viability and invasion in vitro. Oral Surg Oral Med Oral Pathol Oral Radiol 117:234-42 (2014). WB . PubMed: 24439919

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