Anti-Choline Transporter (CHT) (extracellular) Antibody

Na+/Cl-- dependent high affinity choline transporter, SLC5A7
    Cat #: ACT-001
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: h, m, r
    Immunogen
    Peptide (C)DWNQTAYGYPDPK, corresponding to amino acid residues 299-311 of mouse Choline Transporter (Accession Q8BGY9). 4th extracellular loop.
    Accession (Uniprot) Number Q8BGY9
    Gene ID 63993
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Rat, 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, 0.05% NaN3.
    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 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 human SH-5YSY neuroblastoma cell lysate (lanes 1 and 4), rat (lanes 2 and 5) and mouse (lanes 3 and 6) brain lysates:
    1-3. Anti-Choline Transporter (CHT) (extracellular) Antibody (#ACT-001), (1:200).
    4-6. Anti-Choline Transporter (CHT) (extracellular) Antibody, preincubated with the control peptide antigen.
    Immunohistochemistry
    Expression of Choline Transporter in rat striatum
    Immunohistochemical staining of rat frozen brain sections using Anti-Choline Transporter (CHT) (extracellular) Antibody (#ACT-001) (1:200). A. Choline Transporter expression (green) is detected in the matrix (M), but not in the patches (P). B. DAPI (blue) staining provides general cellular staining. C. merge of A and B shows that Choline Transporter appears in a subset of striatal neurons (arrows provide examples).
    Live cell imaging / Immunocytochemistry
    Expression of Choline Transporter in rat PC12 cells
    Immunocytochemical staining of live intact rat pheochromocytoma PC12 cells. A. Cells were stained with Anti-Choline Transporter (CHT) (extracellular) Antibody (#ACT-001), (1:100), followed by goat anti-rabbit-AlexaFluor-594 secondary antibody (red). B. Live view of the cells. C. Merge of the two pictures.
    References
    1. Ribeiro, F.M. et al. (2006) J. neurochem. 97, 1.
    2. Okuda, T. and Haga, T. (2003) Neurochem. Res. 28, 483.
    3. Apparsundaram, S. et al. (2000) Biochem. Biophys. Res. Commun. 276, 862.
    4. Ferguson, S.M. and Blakely, R.S. (2004) Mol. Intervent. 4, 22.
    5. Simon, J.R. and Kuhar, M.G. (1975) Nature 255, 162.
    6. Misawa, H. et al. (2001) Neuroscience 105, 87.
    Scientific background

    In cholinergic neurons, acetylcholine is synthesized from choline and acetyl-coenzyme A (acetyl-CoA). Following depolarization, acetylcholine is released in the synaptic cleft via synaptic vesicles and activates muscarinic and nicotinic receptors located on postsynaptic membranes1,2. Acetylcholine released in the synaptic cleft is rapidly hydrolyzed into choline and acetate. Since choline is not de novo synthesized and thereby only made available through diet uptake, choline is recycled back into the cells in order to regenerate acetylcholine. For this purpose, choline is taken up by the high affinity choline transporter (CHT)1,2.

    CHT has thirteen transmembrane domains, an extracellular N-terminus and an intracellular C-terminus3,4. CHT belongs to the Na+-dependent glucose transporter family (SLC5). The activity of the transporter can be confirmed by its sensitivity to hemicholimium-3 (HC-3) which inhibits the transport of choline with a Ki of 10-100 nM1. The activity of CHT reflects that of neurons1,5 i.e., increased CHT activity indicates an increase in neuronal activity. The increase in the activity of the transporter is due to the increase in the number of transporters and not due to the increase in activity per se. In addition CHT activity is also regulated by second messengers. Phosphorylation of the transporter also seems to determine the activity as it has many consensus phosphorylation sites present in the C-terminal1,2.

    Expression studies reveal that CHT is expressed solely in cholinergic neurons making the transporter a useful marker for detecting these types of neurons5.

    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: 06/11/2018

    Alomone Labs is pleased to offer a highly specific antibody directed against an epitope of mouse CHT. Anti-Choline Transporter (CHT) (extracellular) Antibody (#ACT-001) can be used in western blot analysis and immunocytochemistry applications. It has been designed to recognize CHT from rat, mouse and human samples.

    For research purposes only, not for human use