Anti-EAAT1 (GLAST) (extracellular) Antibody

Excitatory amino acid transporter 1, Sodium-dependent glutamate/aspartate transporter 1, GLAST-1, SLC1A3
    Cat #: AGC-021
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: m, r
    May also work in: h
    Immunogen
    Peptide (C)KQFKTSYEKRSFK, corresponding to amino acids 188-200 of rat EAAT1 (Accession P24942). 2nd extracellular loop.
    Accession (Uniprot) Number P24942
    Gene ID 29483
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Mouse - identical; human 12/13 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 0.4 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, 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:
    1,2. Anti-EAAT1 (GLAST) (extracellular) Antibody (#AGC-021) (1:500).
    3,4. Anti-EAAT1 (GLAST) (extracellular) Antibody, preincubated with the control peptide antigen.
    Immunohistochemistry
    Expression of EAAT1 in rat hippocampus
    Immunohistochemical staining was performed in perfusion-fixed free floating rat brain sections (frozen) using Anti-EAAT1 (GLAST) (extracellular) Antibody (#AGC-021), (1:100). A. EAAT1 staining (green) is particularly intense along the subgranular layer (arrows). B. Staining with mouse anti glial fibrillary acidic protein (red). C. Merged picture confirms presence of densely packed astrocytes along the subgranular layer. DAPI is used as a general cell marker (blue).
    Live cell imaging / Immunocytochemistry
    Expression of EAAT1 in human U-87 MG cells
    Immunocytochemical staining of live intact human U-87 MG glioblastoma cells. A. Extracellular staining of cells using Anti-EAAT1 (GLAST) (extracellular) Antibody (#AGC-021), (1:100) followed by goat anti-rabbit-Alexa-Fluor-594 secondary antibody (red). B. Live image of the cells. C. Merge of A and B.
    References
    1. Tzingounis, A.V. and Wadiche, J.I. (2007) Nat. Rev. Neurosci. 8, 935.
    2. Beart, P.M. and O’Shea, R.D. (2007) Br. J. Pharmacol. 150, 5.
    Scientific background

    L-Glutamate (Glu) is an abundant amino acid that functions as the major excitatory neurotransmitter in the central nervous system. However, excess of Glu in the extracellular synaptic milieu leads to neuronal cell death by a process known as excitotoxicity.

    The extracellular levels of Glu are regulated by a family of high affinity plasma membrane transporters called excitatory amino acid transporters (EAATs) which are responsible for re-uptake of Glu into the cells.1,2 

    The EAAT family includes five members (EAAT1-EAAT5) that are members of the solute carrier family 1 (SLC1) of sodium-dependent transporters that also includes the neutral amino acid transporters ASCT1 and ASCT2.

    The Glu transporters present an unusual topology of eight transmembrane domains with two re-entrant loops and intracellular N- and C- termini. The transporter is likely assembled as a trimer where each monomer is a functional unit capable of binding the Glu substrate.

    The transport of Glu into the cells by the EAAT transporters is coupled to the Na+ and K+ electrochemical gradient as a driving force. Hence, the uptake of Glu is dependent on the co-transport of three Na+ and one H+ ions, and the counter transport of one K+ ion.  

    In addition, to the well documented Glu uptake, the EAAT transporters show a Glu-independent Cl- conductance. The physiological significance of the Cl- current through the EAATs is currently unknown.1,2

    EAAT1 as well as EAAT2, is expressed predominantly in glia cells (hence its original name: glial glutamate transporter or GLAST), while EAAT3EAAT4 and EAAT5 are mostly expressed in neurons.

    As mentioned earlier, EAAT transporters represent the only (significant) mechanism for removal of glutamate from the extracellular fluid and hence are essential for the long-term maintenance of low and non-toxic concentrations of glutamate and the preservation of normal excitatory synaptic transmission.

    In addition, to Glu uptake the glutamate transporters provide glutamate for the synthesis of γ-Aminobutyric acid (GABA), glutathione and protein, suggesting an interactive role between EAATs and cellular metabolism.1,2

    Dysregulation of EAATs activities has been implicated in several neurodegenerative disorders such as Alzheimer’s disease, traumatic brain injury, epilepsy and schizophrenia, suggesting that EAATs can be a useful target for the treatment of these conditions.1,2

    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: 03/12/2018

    Anti-EAAT1 (GLAST) (extracellular) Antibody (#AGC-021) is a highly specific antibody directed against an epitope of rat Excitatory amino acid transporter 1. The antibody can be used in western blot, immunohistochemistry and live cell imaging applications. It has been designed to recognize EAAT1 from rat, human and mouse samples.

    For research purposes only, not for human use