Anti-Dopamine Transporter (DAT) (extracellular) Antibody

Sodium-dependent dopamine transporter, SLC6A3
    Cat #: AMT-003
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: m, r
    Immunogen
    Peptide (C)DAHASNSSDGLGLND, corresponding to amino acids residues 191-205 of rat Dopamine Transporter (Accession P23977). 2nd extracellular loop.
    Accession (Uniprot) Number P23977
    Gene ID 24898
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Mouse - 14/15 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.
    Specificity Not recommended for use with human samples.
    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 -20ºC.
    Preadsorption Control 1 μg peptide per 1 μg antibody.
    Standard quality control of each lot Western blot analysis.
    Applications: ic, ih, lci, wb
    May also work in: ifc, ip
    Western blot
    Western blot analysis of rat (lanes 1 and 3) and mouse (lanes 2 and 4) brain membranes and rat pheochromocytoma PC12 (lanes 3 and 6) cell lysates:
    1-3. Anti-Dopamine Transporter (DAT) (extracellular) Antibody (#AMT-003), (1:200).
    4-6. Anti-Dopamine Transporter (DAT) (extracellular) Antibody, preincubated with the control peptide antigen.
    Immunohistochemistry
    Mouse coronal brain sections (1:200) (Hong, J. et al. (2015) Cell Death Dis. 6, e1832.).
    Live cell imaging / Immunocytochemistry
    Expression of DAT in rat PC12 cells
    Immunocytochemical staining of live intact rat pheochromocytoma PC12 cells. A. Cells stained with Anti-Dopamine Transporter (DAT) (extracellular) Antibody (#AMT-003), (1:100), followed by goat anti-rabbit-AlexaFluor- 594 secondary antibody (red). B. Live view of the cells.
    References
    1. Robertson, S.D. et al. (2009) Mol. Neurobiol. 39, 73.
    2. Gainetdinov, R.R. (2008) Nauyn-Schiedeberg’s Arch. Pharmacol. 377, 301.
    3. Torres, G.E. and Amara, S.G. (2007) Curr. Opin. Neurobiol. 17, 304.
    4. Ciliax, B.J. et al. (1995) J. Neurosci. 15, 1714.
    5. Uhl, G.R. (2003) Mov. Disord. 18, S71.
    Scientific background

    The Na+/Cl- transporter family, SLC6, includes DAT a monoamine transporter, important for regulating extracellular levels of dopamine. It does so by uptaking dopamine from the synaptic cleft via the co-transport of Na+ and Cl- down their electrochemical gradient1. The removal of dopamine by DAT remains the most important means to control the extracellular lifetime of the neurotransmitter, and notably the ending of dopaminergic neurotransmission2. Transporters for serotonin, norepinephrine, GABA and glycine also belong to this family3.

    These receptors have 12 transmembrane spanning domains and intracellular N- and C-terminal domains. DAT is also subjected to post translational modifications such phosphorylation, important for its regulation1. Also, it possesses a large extracellular domain which undergoes N-glycosylation, important for the proper targeting of the transporter to the plasma membrane1. DAT is expressed in dopaminergic cell bodies and terminals and can therefore serve as a marker for these neurons4. DAT is also expressed in the retina, gastrointestinal tract, lung, kidney, pancreas and lymphocytes2.

    DAT plays an important role in movement as well as reward, learning and memory5. Its malfunction which leads to dopaminergic dysregulation has been associated with ADHD, schizophrenia, as well as Parkinson’s disease2. DAT is the main target for cocaine and amphetamine and methamphetamine psychostimulants, which mainly increase locomotor activities2.

    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
    Image & Title:
    Expression of Dopamine Transporter in mouse brain.Western blot analysis of mouse midbrain lysates using Anti-Dopamine Transporter (DAT) (extracellular) Antibody (#AMT-003) (upper panel). DAT protein levels from mice exposed to different drugs are shown. Immunohistochemical staining of mouse brain sections (lower panel). DAT staining (red) in substantia nigra is detected in dopaminergic neurons.Adapted from Hong, J. et al. (2015) Cell Death Dis. 6, e1832. with permission of Nature Publishing Group.
    Last update: 01/11/2018

    Anti-Dopamine Transporter (DAT) (extracellular) Antibody (#AMT-003) is a highly specific antibody directed against an epitope of the rat protein. The antibody can be used in western blot, immunohistochemistry, and live cell imaging applications. It has been designed to recognize DAT from mouse and rat samples. The antibody is unlikely to recognize DAT from human samples.

    For research purposes only, not for human use
    Citations
    Western blot citations
    1. Mouse brain lysate (1:200).
      Hong, J. et al. (2015) Cell Death Dis. 6, e1832.
    Immunohistochemistry citations
    1. Mouse coronal brain sections (1:200).
      Hong, J. et al. (2015) Cell Death Dis. 6, e1832.
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