Anti-HCN2 Antibody

Hyperpolarization-activated cyclic nucleotide-gated potassium channel 2, BCNG2, HAC-1
    Cat #: APC-030
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: h, m, r
    Immunogen
    Peptide (C)EEAGPAGEPRGSQAS, corresponding to amino acid residues 147-161 of human HCN2 (Accession Q9UL51). Intracellular, N-terminus.
    Accession (Uniprot) Number Q9UL51
    Gene ID 114244
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Rat, mouse - 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 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, ih, ip, wb
    May also work in: ifc
    Western blot
    Western blot analysis of rat brain membranes:
    1. Anti-HCN2 Antibody (#APC-030), (1:200).
    2. Anti-HCN2 Antibody, preincubated with the control peptide antigen.
    Immunoprecipitation
    Rat thalamus lysate (Whitacker, G.M. et al. (2007) J. Biol. Chem. 282, 22900.).
    Immunohistochemistry
    Expression of HCN2 in rat cerebellum
    Immunochistochemical staining of rat cerebellum frozen sections using Anti-HCN2 Antibody (#APC-030 ). A. HCN2 (red) appears in Purkinje cells (arrows). B. Staining of astrocytes with mouse anti-glial fibrillary acidic protein (GFAP, green demonstrates the restriction of HCN2 to neuronal cell bodies. C. Confocal merge of HCN2 and GFAP images demonstrates the respective localization of these proteins.
    Expression of HCN2 in mouse hypothalamus
    Immunohistochemical staining of mouse hypothalamus using Anti-HCN2 Antibody (#APC-030). A. HCN2 (red) appears in cells of the paraventricular nucleus (PVN, arrows). B. Staining of paraventricular nerve cells with mouse anti-calcium binding protein (CBD28k, green). C. Confocal merge of HCN2 and CBD28k demonstrates some co-localization. V = Third ventricle.
    Colocalization of HCN4 and HCN2 in mouse thalamus
    Immunohistochemical staining of mouse thalamus frozen section using Guinea pig Anti-HCN4 Antibody (#AGP-004) and rabbit Anti-HCN2 Antibody (#APC-030). A. Staining of HCN4 (green) appears in the ventral posterior thalamic nucleus (VPL). B. In the same section as in A, staining of HCN2 (red) appears in the ventral posterior thalamic nucleus (VPL) and also in the reticular thalamic nucleus (RT). The area between these thalamic nuclei (star) is white matter and neither protein is expressed in that region. C. Merged images of A and B.
    Immunocytochemistry
    Expression of HCN2 in rat DRG primary culture
    Immunocytochemical staining of paraformaldehyde-fixed and permeabilized rat dorsal root ganglion (DRG) primary culture using Anti-HCN2 Antibody (#APC-030), (1:100), (green). Cells were stained with Anti-HCN2 Antibody followed by goat anti-rabbit-AlexaFluor-488 secondary antibody. Nuclear staining of cells using the cell-permeable DNA dye Hoechst 33342 (blue).
    References
    1. Much, B. et al. (2003) J. Biol. Chem278, 43781.
    2. Notomi, T. et al. (2004) J. Comp. Neurol471, 241.
    3. Hofmann, F. et al. (2005) Pharmacol. Rev. 57, 455.
    Scientific background

    Hyperpolarization-activated cation currents (Ih) appear in the heart and the brain having crucial role in controlling electrical pacemaker activity, contributing to biological processes such as heartbeat, sleep-wake cycle and synaptic plasticity.1,2

    The Ih currents are generated by the hyperpolarization-activated cyclic nucleotide-gated channel family (HCN), which comprises four homologous members, named HCN1-4. Each HCN subunit consist of six transmembrane domains (TM), a pore region between TM5-TM6 and a binding domain to cyclic nucleotides (CNBD) in the cytoplasmic C-terminus.2

    The HCN subunits can form functional homomers and can also co-assemble into functional heteromers.2

    The channels are closely related to each other and share homology of about 60%. However, they are diverging from each other in their cytoplasmic N- and the C-terminus. The channels HCN1-4 mainly differ from each other with regard to their speed of activation and the extent by which they are modulated by cAMP. HCN1 is the fastest channel, followed by HCN2, HCN3 and HCN4.2,3

    HCN2 is the most abundant neuronal channel and is found almost ubiquitously in the brain.3

    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:


    Knockout validation of Anti-HCN2 Antibody in mouse brain.Immunohistochemical staining of mouse coronal brain sections using Anti-HCN2 Antibody (#APC-030). HCN2 staining (red) is broadly detected across the coronal section. Lack of HCN2 staining is observed in HCN2-/- mice. Adapted from Hammelmann, V. et al. (2011) PLoS ONE 6, e17078.

    Last update: 22/11/2018

    Anti-HCN2 Antibody (#APC-030) is a highly specific antibody directed against an epitope of the human protein. The antibody can be used in western blot, immunoprecipitation, immunohistochemistry, and immunocytochemistry applications. It has been designed to recognize HCN2 from human, rat, and mouse samples.

    For research purposes only, not for human use
    Citations
    KO validation citations
    1. Immunohistochemical staining of mouse brain sections. Also tested in HCN2-/- mice.
      Hammelmann, V. et al. (2011) PLoS ONE 6, e17078.
    Western blot citations
    1. Human sinoatrial node lysate (1:500).
      Li, N. et al. (2015) Circ. Arrhythm. Electrophysiol. 8, 1219.
    Immunoprecipitation citations
    1. Rat thalamus lysate.
      Whitacker, G.M. et al. (2007) J. Biol. Chem. 282, 22900.
    Immunohistochemistry citations
    1. Rat diaphragm sections (1:25).
      Negrini, D. et al. (2016) Am. J. Physiol. 311, H892.
    2. Human sinoatrial node sections (1:100).
      Li, N. et al. (2015) Circ. Arrhythm. Electrophysiol. 8, 1219.
    3. Mouse vestibular ganglia.
      Horwitz, G.C. et al. (2014) J. Gen. Physiol. 143, 481.
    4. Mouse brain sections. Also tested in HCN2-/- mice.
      Hammelmann, V. et al. (2011) PLoS ONE 6, e17078.

    More product citations
    1. Cao Ehlker, Xet al. (2013) J. Biol. Chem. 288, 7580.
    2. Nakashima, N. et al. (2013) J. Physiol. 591, 1749.
    3. Rusznak, Z. et al. (2013) Eur. J. Neurosci. 37, 876.
    4. Weisbrod, D. et al. (2013) Proc. Natl. Acad. Sci. U.S.A. 110, E1685.
    5. Ramakrishnan, N.Aet al. (2012) J. Biol. Chem. 287, 37628.
    6. Doan, T.Net al. (2004) J. Neurosci. 24, 3335.
    7. Jiang, Met al. (2004) Circulation 109, 1783.
    8. Macri, V. and Accili, A.A. (2004) J. Biol. Chem. 279, 16832.
    9. Much, Bet al. (2003) J. Biol. Chem. 278, 43781.
    10. Han, Wet al. (2002) Circ. Res91, 790.
    11. Moroni Aet al. (2001) J. Biol. Chem276, 29233.
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