Anti-TRPM8 (extracellular) Antibody

TRPP8, LTrpC6, Transient receptor potential cation channel subfamily M member 8
    Cat #: ACC-049
  • Lyophilized Powder
  • Antigen Incl.
  • Shipped at Room Temp.
  • Type: Polyclonal
    Source: Rabbit
    Reactivity: h, m, r
    Immunogen
    Peptide SDVDGTTYDFAHC, corresponding to amino acid residues 917-929 of human TRPM8 (Accession Q7Z2W7). 3rd extracellular loop.
    Accession (Uniprot) Number Q7Z2W7
    Gene ID 79054
    Peptide confirmation Confirmed by amino acid analysis and mass spectrometry.
    Homology Canis - identical; bovine - 12/13 amino acid residues identical; rat, mouse - 11/13 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 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, lci, n, wb
    May also work in: ifc
    Western blot
    Western blot analysis of prostate carcinoma cell lines; DU145 (lanes 1, 3), LNCaP (lanes 2, 4) and mouse-TRPM8 transfected HEK-293 (lanes 5, 6) cell lysates:
    1,2,5. Anti-TRPM8 (extracellular) Antibody (#ACC-049), (1:200).
    3,4,6. Anti-TRPM8 (extracellular) Antibody, preincubated with the control peptide antigen.
    Immunoprecipitation
    Transfected HEK-293 cells (Zeevi, D.A. et al. (2010) J. Cell Sci. 123, 3112.).
    Immunohistochemistry
    Expression of TRPM8 in rat DRG
    Immunohistochemical staining of rat dorsal root ganglion (DRG) frozen sections using Anti-TRPM8 (extracellular) Antibody (#ACC-049), (1:100). TRPM8 is expressed in DRG neurons. Hoechst 33342 is used as the counterstain.
    Immuno-colocalization of TRPM8 and TrkA in rat DRG
    Immunohistochemical staining of perfusion-fixed frozen rat dorsal root ganglia (DRG) sections using Anti-TrkA (extracellular)-ATTO-633 Antibody (#ANT-018-FR), (1:60) and Anti-TRPM8 (extracellular) Antibody (#ACC-049), (1:300). A. TRPM8 labeling followed by goat-anti-rabbit-Alexa-488 (green). B. The same section was then labeled for TrkA (purple).  C. Merge of A and B demonstrates co-localization of TRPM8 and TrkA in rat DRG (arrows). Cell nuclei were stained with DAPI (blue).
    Immunocytochemistry
    Human prostate cancer DU145 cells (1:50) (Wang, Y. et al. (2012) Pathol. Oncol. Res18, 903.).
    Neutralization
    Human, mouse and rat TRPM8 expressed in CHO cells, and rat DRGs (Miller, S. et al. (2014) PLoS ONE 9, e107151).
    Live cell imaging / Immunocytochemistry
    Expression of TRPM8 in LNCaP prostate carcinoma cell line
    Immunocytochemical staining of LNCaP cells with Anti-TRPM8 (extracellular) Antibody (#ACC-049). A. Extracellular staining of intact LNCaP cells with Anti-TRPM8 (extracellular) Antibody (1:100), followed by goat anti-rabbit-AlexaFluor-550 (red), (x100). B. Intracellular staining of LNCaP cells with Anti-TRPM8 (extracellular) Antibody (1:1000), followed by goat anti-rabbit-AlexaFluor-488 (green), (x100).
    Expression of TRPM8 in rat DRG cells
    Immunocytochemical staining of intact living rat root dorsal root ganglion (DRG) cells. A. Intracellular staining of cells with Anti-TRPM8 (extracellular) Antibody (#ACC-049), (1:500) followed by goat anti-rabbit-AlexaFluor-555 secondary antibody. B. Extracellular staining of live cells with Anti-TRPM8 (extracellular) Antibody (1:50) followed by goat anti-rabbit-AlexaFluor-555 secondary antibody. The cell-permeable dye Hoechst 33342 (blue) was used for nuclear staining.
    References
    1. Fleig, A. et al. (2004) Trends. Pharmacol. Sci. 25, 633.
    2. Brauchi, S. et al. (2004) Proc. Natl. Acad. Sci.  101, 15494.
    3. McKemy, D.D. et al. (2002) Nature 416, 52.
    4. Peier, A.M. et al. (2002) Cell 108, 705.
    5. Dragoni, I. et al. (2006) J. Biol. Chem. 281, 37353.
    6. Zhang, L. et al. (2006) Endocr. Relat. Cancer 13, 27.
    Scientific background

    The mammalian melastatin-related transient receptor potential (TRPM) is a subfamily of the TRP family. The family was named after the first member, melastatin (TRPM1), whose gene was identified in metastatic and benign melanomas.1 The TRPM proteins share structural homology with other members of the TRP superfamily channels; six putative transmembrane domains, and cytoplasmic N-terminus and C-terminus. However, due to their long N-terminus and C-terminus they were also named the long TRP channel family.1

    The TRPM family consists of eight members designated as TRPM1-8 that can be further divided into four pairs: TRPM1 and TRPM3; TRPM2 and TRPM8; TRPM4 and TRPM5; and TRPM6 and TRPM7.1

    The TRPM8 channel is the cold and menthol receptor activated by either cold temperatures(~28°C) or menthol.2

    TRPM8 is expressed in dorsal root ganglia (DRGs) where about 5%-10% of the small diameter DRG neurons express the channel.3,4 In DRGs, TRPM8 expressing neurons do not express the TRPV1 channel.5 Overexpression of TRPM8 was found in prostate cancer cells. However, the physiological and pathological roles that these cells play is still elusive.6 In prostate, it was suggested that TRPM8 might play a possible role in the progression of cancer to the metastatic stage.6

    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:

    Anti-TRPM8 (extracellular) Antibody
    Expression of TRPM8 in rat tail artery astrocytes.Immunocytochemical staining of rat tail artery astrocytes using Anti-TRPM8 (extracellular) Antibody (#ACC-049). TRPM8 staining is shown in green, nuclei are in blue. Omission of the antibody (lower panel) abolishes TRPM8 staining.Adapted from Johnson, C.D. et al. (2009) Am. J. Physiol. 296, H1868. with permission of the American Physiological Society.

    Last update: 08/11/2018

    Anti-TRPM8 (extracellular) Antibody (#ACC-049) is a highly specific antibody directed against an epitope of the human protein. The antibody can be used in western blot, immunoprecipitation, immunohistochemistry, immunocytochemistry, and neutralization applications. It has been designed to recognize TRPM8 from human, rat, and mouse samples.

    For research purposes only, not for human use
    Citations
    Published figures using this product
    Anti-TRPM8 (extracellular) Antibody specifically blocks TRPM8.
    A. Human TRPM8 cold activation is blocked by 2.5 μM of Anti-TRPM8 (extracellular) Antibody (#ACC-049). B. TRPM8 iciclin activation is blocked by 2.5 μM of the same antibody. M8-B is a specific TRPM8 antagonist. C. Anti-TRPM8 (extracellular) Antibody does not block cold activation of TRPA1 and heat activation of TRPV1 (D). AMG9090 and AMG6541 are specific TRPA1 and TRPV1 antagonist.
    Adapted from Miller, S. et al. (2014) with kind permission of Dr. Gavva, N.R. Department of Neuroscience, Amgen Inc., California, U.S.A.
    Western blot citations
    1. Transfected HEK293 cell lysate (1:500).
      Gkika, D. et al. (2015) J. Cell Biol. 208, 89.
    2. Rat F11 dorsal root ganglion hybridoma cells lysate (1:400).
      Toro, C.A. et al. (2015) J. Neurosci. 35, 571.
    Immunoprecipitation citations
    1. Transfected HEK 293 cells (7 µg/ml).
      Vinuela-Fernandez, I. et al. (2014) Neuropharmocolgy 79, 136.
    2. Transfected HEK 293 cells.
      Zeevi, D.A. et al. (2010) J. Cell Sci. 123, 3112.
    Immunohistochemistry citations
    1. Rat DRG sections (1:300).
      Vinuela-Fernandez, I. et al. (2014) Neuropharmocolgy 79, 136.
    Immunocytochemistry citations
    1. Rat tail artery VSMC (1:200).
      Melanaphy, D. et al. (2016) Am. J. Physiol. 311, H1416.
    2. Rat F11 dorsal root ganglion hybridoma cells (1:400).
      Toro, C.A. et al. (2015) J. Neurosci. 35, 571.
    3. Human prostate cancer DU145 cells (1:50).
      Wang, Y. et al. (2012) Pathol. Oncol. Res18, 903.
    4. Rat tail artery astrocytes.
      Johnson, C.D. et al. (2009) Am. J. Physiol. 296, H1868.
    Neutralization citations
    1. Human, mouse and rat TRPM8 expressed in CHO cells, and rat DRGs.
      Miller, S. et al. (2014) PLoS ONE 9, e107151.
    More product citations
    1. Ma, S. et al. (2012) J. Mol. Cell Biol. 4, 88.
    2. El Karim, I.A. et al. (2011) J. Endod. 37, 473.
    3. El Karim, I.A. et al. (2011) Pain 152, 2211.
    4. Harrington, A.M. et al. (2011) Pain 152, 1459.
    5. Martinez Lopez, P. et al. (2011) J. Cell Physiol. 226, 1620.
    6. Johnson, C.D. et al. (2009) Am. J. Physiol. 296, H1868.
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