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Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody

Protease-activated receptor-4, PAR-4, Coagulation factor II receptor-like 3, Thrombin receptor-like 3

Cat #: APR-034-F
Alternative Name Protease-activated receptor-4, PAR-4, Coagulation factor II receptor-like 3, Thrombin receptor-like 3
  • KO Validated
  • Lyophilized Powder yes
    Type: Polyclonal
    Host: Rabbit
    Reactivity: h, m, r
    Immunogen
    • Peptide (C)HLRGQRWPFGEAA(S)R, corresponding to amino acid residues 136-150 of human PAR-4 (Accession Q96RI0). Cys 149 was replaced with Ser. 1st extracellular loop.
    Accession (Uniprot) Number Q96RI0
    Gene ID 9002
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Rat, mouse - identical.
    RRID AB_2040087.
    Purity Affinity purified on immobilized antigen.
    Form Lyophilized powder. Reconstituted antibody contains phosphate buffered saline (PBS), pH 7.4, 1% BSA, 0.05% NaN3.
    Isotype Rabbit IgG.
    Label Fluorescein isothiocyanate (FITC).
    Storage before reconstitution The antibody ships as a lyophilized powder at room temperature. Upon arrival, it should be stored at -20°C.
    Reconstitution 50 µl double distilled water (DDW).
    Antibody concentration after reconstitution 1 mg/ml.
    Storage after reconstitution The reconstituted solution can be stored at 4°C, protected from the light, for up to 1 week. For longer periods, small aliquots should be stored at -20°C. Avoid multiple freezing and thawing. Centrifuge all antibody preparations before use (10000 × g 5 min).
    Standard quality control of each lot Western blot analysis (unlabeled antibody, #APR-034), and direct flow cytometry (labeled antibody).
    Applications: fc
    May also work in: ic*, lci*
    Direct flow cytometry
    • Cell surface detection of PAR-4 in live intact human MEG-01 megakaryoblastic leukemia cells:
      Cell surface detection of PAR-4 in live intact human MEG-01 megakaryoblastic leukemia cells:
      ___ Cells alone.
      ___ Cells + rabbit IgG isotype control-FITC.
      ___ Cells + Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody (#APR-034-F), 5µg.
    • Cell surface detection of PAR-4 in live intact HL-60 (human promyelocytic leukemia) cell line:
      Cell surface detection of PAR-4 in live intact HL-60 (human promyelocytic leukemia) cell line:
      ___ Unstained cells.
      ___ Cells + Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody (#APR-034-F).
    • Cell surface detection of PAR-4 in human neutrophil-like differentiated promyelocytic leukemia HL-60 cells
      Cell surface detection of PAR-4 in human neutrophil-like differentiated promyelocytic leukemia HL-60 cells
      HL-60 cells were induced to differentiate into a mature neutrophil-like phenotype by incubation with dimethyl sulfoxide (DMSO) (1.25%) for three days. The neutrophil phenotype was confirmed by staining the cells with anti-CD11b-PE antibody. Double staining with Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody (#APR-034-F), (1:40), shows that most differentiated cells express both markers (upper right panel).
    • The control antigen is not suitable for this application.
    References
    1. MacFarlane, S.R. et al. (2001) Pharmacol. Rev. 53, 245.
    2. Hollenberg, M.D. et al. (2002) Pharmacol. Rev. 54, 203.
    3. Ossovskaya, V.S. et al. (2004) Physiol. Rev. 84, 579.
    Scientific background

    Protease-activated receptor 4 (PAR-4) belongs to a family of four G protein-coupled receptors (PAR1-4) that are activated as a result of proteolytic cleavage by certain serine proteases, hence their name. In this novel modality of activation, a specific protease cleaves the PAR receptor within a defined sequence in its extracellular N-terminal domain. This results in the creation of a new N-terminal tethered ligand, which subsequently binds to a site in the second extracellular loop of the same receptor. This binding results in the coupling of the receptor to G proteins and in the activation of several signal transduction pathways.1-3

    Different PARs are activated by different proteases. Hence, PAR-4 is activated by both thrombin and trypsin whereas PAR-1 and PAR-3 are activated only by thrombin and PAR-2 is activated only by trypsin.1-3 PAR-4 can be also cleaved and activated by other proteases such as cathepsin G.

    The intracellular signaling mechanisms mediated by PAR-4 activation are not completely elucidated but they involve calcium mobilization downstream of phospholipase Cβ through the Gαq pathway.1-3

    Tissue distribution of PAR-4 is very broad with the highest expression levels found in lung, testis, pancreas and small intestine. In addition, PAR-4 expression was observed in platelets, megakaryocytes and leukocytes. Studies with platelets derived form PAR-4 knockout mice have established an essential role for PAR-4 in thrombin-induced platelet activation.
    PAR-4 is likely involved in other physiological functions such as regulation of gastrointestinal motility and regulation of vascular endothelial cell function.1-3

    Application key:

    CBE- Cell-based ELISA, FC- Flow cytometry, ICC- Immunocytochemistry, IE- Indirect ELISA, IF- Immunofluorescence, IFC- Indirect flow cytometry, IHC- Immunohistochemistry, IP- Immunoprecipitation, LCI- Live cell imaging, N- Neutralization, WB- Western blot

    Species reactivity key:

    H- Human, M- Mouse, R- Rat
    Image & Title:

    Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody
    Expression of PAR-4 on mouse platelets.Direct flow cytometry analysis of PAR-4 expression in wild type (black lines), PAR3-/- (grey line), and PAR4-/- (shaded) mice platelets using Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody (#APR-034-F). Note the lack of PAR-4 detection in PAR4-/- mice.Adapted from Arachiche, A. et al. (2013) PLoS ONE 8, e55740. with permission of PLoS.

    Last update: 02/03/2021

    Anti-PAR4 (F2RL3) (extracellular) Antibody (#APR-034) is a highly specific antibody directed against an extracellular epitope of human protease-activated receptor-4. The antibody can be used in western blot analysis, immunohistochemical, immunocytochemical and flow cytometry applications. It has been designed to recognized PAR-4 from human, mouse and rat samples.

    Anti-PAR4 (F2RL3) (extracellular)-FITC Antibody (#APR-034-F) is directly conjugated to fluorescein isothiocyanate (FITC). This labeled antibody can be used in immunofluorescent applications such as direct live cell flow cytometry.

    For research purposes only, not for human use

    Applications

    Specifications

    Scientific Background

    Citations

    Citations
    KO validation citations
    1. Direct flow cytometry analysis of mouse platelets. Tested in PAR4-/- mice platelets.
      Arachiche, A. et al. (2013) PLoS ONE 8, e55740.
    Direct flow cytometry citations
    1. Human macrophage U937 cells.
      Mahajan-Thakur, S. et al. (2014) J. Leukoc. Biol. 96, 611.
    2. Mouse platelets. Also tested in PAR4-/- mice platelets.
      Arachiche, A. et al. (2013) PLoS ONE 8, e55740.
    3. Human peripheral blood mononuclear cells (PBMCs).
      Nieuwenhuizen, L. et al. (2013) Scand. J. Immunol. 77, 339.
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