Anti-TrkA (extracellular) Antibody

High affinity nerve growth factor receptor, Neurotrophic tyrosine kinase receptor type 1, NTRK1
    Cat #: ANT-018
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: h, m, r
    Immunogen
    Peptide (C)ETMRHG(S)LRLNQPTH, corresponding to amino acid residues 342-356 of rat TrkA (Accession P35739). 2nd extracellular immunoglobulin-like domain.
    Accession (Uniprot) Number P35739
    Gene ID 59109
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Mouse - identical; human - 13/14 amino acid residues identical.
    Purity Affinity purified on immobilized antigen.
    Formulation Lyophilized powder. Reconstituted antibody contains phosphate buffered saline (PBS), pH 7.4, 1% BSA, 0.025% NaN3.
    Storage before reconstitution The antibody ships as a lyophilized powder at room temperature. Upon arrival, it should be stored at -20°C.
    Reconstitution 25 µl, 50 μl or 0.2 ml double distilled water (DDW), depending on the sample size.
    Antibody concentration after reconstitution 0.8 mg/ml.
    Storage after reconstitution The reconstituted solution can be stored at 4°C 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 x g 5 min).
    Control antigen storage before reconstitution Lyophilized powder can be stored intact at room temperature for 2 weeks. For longer periods, it should be stored at -20°C.
    Control antigen reconstitution 100 µl double distilled water (DDW).
    Control antigen storage after reconstitution -20°C.
    Preadsorption Control 1 μg peptide per 1 μg antibody.
    Standard quality control of each lot Western blot analysis.
    Applications: ic, ifc, ih, lci, wb
    May also work in: ip
    Western blot
    Western blot analysis of rat (lanes 1 and 3) and mouse (lanes 2 and 4) brain lysates:
    1,2. Anti-TrkA (extracellular) Antibody (#ANT-018), (1:200).
    3,4. Anti-TrkA (extracellular) Antibody, preincubated with the control peptide antigen.
    Immunohistochemistry
    Expression of TrkA in rat DRG
    Immunohistochemical staining of rat dorsal root ganglia (DRG) frozen sections using Anti-TrkA (extracellular) Antibody (#ANT-018), (1:100). TrkA (red staining) is expressed in DRG neurons and in satellite microglia (arrows). Hoechst 33342 is used as the counterstain.
    Expression of TrkA in rat brain hippocampal dentate gyrus
    Immunohistochemical staining of immersion-fixed, free floating rat brain frozen sections. A. Brain sections were stained using Anti-TrkA (extracellular) Antibody (#ANT-018), (1:1000), (green staining). B. The same section was also stained for glial fibrillary acidic protein (GFAP) (red and counterstained blue). C. Overlay of A and B demonstrates co-localization of TrkA and GFAP in dentate gyrus astrocytes.
    Indirect flow cytometry
    Indirect flow cytometry analysis of live intact Jurkat (human T cell leukemia) cell line:
    ___ Cells + goat-anti-rabbit-Cy5.
    ___ Cells + Anti-TrkA (extracellular) Antibody (#ANT-018), (1:25) + goat-anti-rabbit-Cy5.
    The control antigen is not suitable for this application.
    Live cell imaging / Immunocytochemistry
    Expression of TrkA in live intact rat PC12 cells
    Immunocytochemical staining of live intact rat PC12 cells. A. Cells were stained with Anti-TrkA (extracellular) Antibody (#ANT-018) (1:50), followed by goat anti-rabbit-AlexaFluor-494 secondary antibody (red). B. Cell nuclei were visualized with the membrane-permeable DNA dye Hoechst 33342 (blue staining). C. Live view of the cells.
    References
    1. Webster, N.J.G. and Pirrung, M.C. (2008) BMC Neurosci. 9, S1.
    2. Quartu, M. et al. (2003) Int. J. Dev. Neurosci. 21, 309.
    3. Aoki, C. et al. (2000) J. Neurosci. Res. 59, 454.
    4. Yan, Q. et al. (1997) J. Comp. Neurol. 378, 135.
    5. Crowley, C. et al. (1994) Cell 76, 1001.
    6. Smeyne, R.J. et al. (1994) Nature 368, 246.
    7. Fagan, A.M. et al. (1997) J. Neurosci. 17, 7644.
    8. Patapoutian, A. and Reichardt, L.F. (2001) Curr. Opin. Neurobiol. 11, 272.
    9. Schecterson, L.C. and Bothwell, M. (2010) Develop. Neurobiol. 70, 332.
    10. Grimes, M.L. et al. (1996) J. Neurosci. 16, 7950.
    Scientific background

    NGF was the first neurotrophin to be discovered more than 50 years ago. Three other proteins have been added to the list: BDNFNT-3 and NT-4 (NT-4/5). These neurotrophins bind two groups of receptors. The p75NTR receptor is common to all four neurotrophins and is a member of the tumor necrosis factor receptor family. The tropomyosin-related kinase (TrK) receptors are receptor tyrosine kinases (RTKs) and three receptors form this family: TrkA, TrkB, and TrkC1.

    As mentioned, the p75NTR receptor binds to all neurotrophins with similar affinities while the Trk receptors display selectivity for the neurotrophins. TrkA is activated by NGF binding, TrkB by that of BDNF and NT-4, while TrkC is stimulated by the binding of NT-31.

    All three Trk receptors are highly expressed in the mammalian brain in very distinct regions and are also expressed in the peripheral nervous system2-4. Cholinergic neurons in the basal forebrain exclusively express TrkA. All three Trk receptors are expressed in the hippocampus. Motor and sensory neurons in the peripheral nervous system express Trk receptors. Interestingly, while Trk receptors are not essential for development, knockout mice die shortly after birth. Indeed, TrkA-deficient mice demonstrate a significant decrease in cholinergic and sympathetic neurons in the trigeminal, superior cervical and dorsal root ganglia5-7.

    Trk receptors are type I membrane proteins (i.e. span the membrane with one transmembrane region) and have many motifs in the extracellular region, including cell-adhesion domains, three tandem leucine rich motifs flanked by two clusters of cysteines. In the membrane proximal region of the receptor there are also two immunoglobulin-like domains8. The intracellular region of the protein contains the kinase domain, responsible for activating downstream effectors as well as many adaptor site regions important for the binding/interaction of signaling molecules.

    The binding of neurotrophins to Trk receptors promotes receptor dimerization resulting in kinase activation. Activated Trk receptors then phosphorylate a cascade of signaling molecules including the Ras/ERK, PI3K/Akt pathways and PLC-γ1. Activated Trk receptors also create internal docking sites for other signaling adaptor proteins to bind8.

    Splice variants of TrkA, TrkB and TrkC have been observed. These splice isoforms are mainly effected in the tyrosine kinase domain of the receptor lying in the cytoplasm. Endocytosis is an important signaling trait of Trk receptors. Following neurotrophin binding to the Trk receptor, the receptor complex is then internalized via endocytosis in order to terminate signaling. However, in the axonal compartment of neurons, the internalization process of the neurotrophin complexed to the receptor is part of the signaling process and is important for activating transcription processes in the nucleus9,10.

    Application key:

    CBE- Cell-based ELISA, FC- Flow cytometry, ICC- Immunocytochemistry, IE- Indirect ELISA, 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
    Last update: 29/11/2018

    Anti-TrkA (extracellular) Antibody (#ANT-018) is a highly specific antibody directed against an epitope of the rat protein. The antibody can be used in western blot, immunohistochemistry, live cell imaging, and indirect flow cytometry applications. It has been designed to recognize TrkA from mouse, rat, and human samples.

    For research purposes only, not for human use
    Citations
    Western blot citations
    1. Rat brain lysate.
      Manohar, S. et al. (2016) Mol. Cell. Neurosci. 75, 101.
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