ASIC3 Channel Overexpressed Membrane Fractions

A Positive Control for Detecting ASIC3 Channels in Western Blot Analysis
Cat #: LX-100
  • Lyophilized Powder
  • Accession (Uniprot) Number NM_173135
    Gene ID 286920
    Origin Rat ASIC3 cDNA (NM_173135) was transcribed in vitro to produce full length mRNA. ASIC3 mRNA was injected into Xenopus oocytes and protein expression was verified by detection of pH evoked APETx2 (#RTA-100) sensitive ionic current. Membrane fraction was prepared from oocyte population homogenate and lyophilized. Non-injected oocytes underwent the same procedure excluding mRNA injection.
    Storage before reconstitution Lyophilized injected and non-injected membrane fractions can be stored at room temperature for 2 weeks. For longer periods, they should be stored at -20°C.
    Reconstitution Work on ice. Gently reconstitute injected and non-injected membrane fractions in 100 μl DDW (total protein concentration is 0.2 μg/μl). Centrifuge all product preparations before use (10000 x g 5 min).
    Storage after reconstitution Up to 2 weeks at -20°C. Avoid repeated thawing and freezing.
      • Western blot analysis of ASIC3 Channel Overexpressed Membrane Fractions (#LX-100) obtained from Xenopus oocyte membrane fractions expressing rat ASIC3 channels (lanes 2 and 4), and non-injected oocyte lysates (lanes 1 and 3):
        1, 2. Anti-ASIC3 Antibody (#ASC-018), (1:200).
        3, 4. Anti-ASIC3 Antibody, preincubated with the control peptide antigen.
    Note

    Lyophilized membrane fractions were reconstituted in 100 μl DDW (see below). 100 μl of Sample Buffer X2 was added (total protein concentration was 0.1 μg/μl). Boiled samples were loaded onto 10% acrylamide gel (10-40 μl/lane) and resolved using SDS-PAGE gel, followed by western blot analysis using Anti-ASIC3 Antibody (#ASC-018) following product instructions.

    This product includes:
    1 x 0.1 ml lyophilized ASIC3 channel overexpressed membrane lysate
    1 x 0.1 ml lyophilized non-injected Xenopus oocyte membrane lysate

      • ASIC3 is a member of a family of Na+ channels that are activated by external protons. The family includes another three members ASIC1ASIC2 and ASIC4. The ASICs are in fact part of a larger superfamily termed degenerin/epithelial Na+ channels (DEG/ENaC) and share with it the same basic characteristics: two transmembrane spanning domains, a large extracellular domain and short intracellular N- and C-termini.

        The functional channel is composed of 4 subunits that can be assembled as homo or heterotetramers with the other ASIC subunits.

        A drop in external pH opens the channel resulting in an inward cation current that is quickly inactivated even in the continuous presence of protons in the medium, although a small residual current may persist.

        Several lines of evidence indicate that ASIC3 may function as a pain sensor. First, it is specifically located in dorsal root ganglia (DRG) neurons and in nociceptors that are involved in pain detection and transmission. In addition, in conditions such as inflammation and cardiac ischemia that include tissue acidosis and thus pain, ASIC3 currents have been detected. Finally, ASIC3 deficient mice display altered sensitivity to high intensity pain produced by heat or acid.

        Interestingly, while in rats ASIC3 is expressed almost exclusively in DRG neurons, in humans its expression is more widespread suggesting a more extensive role in human nociception.

    Last update: 08/04/2019

    ASIC3 Channel Overexpressed Membrane Fractions (#LX-100) are Xenopus oocyte membrane fractions overexpressing the ASIC3 channel.* ASIC3 Channel Overexpressed Membrane Fractions are your positive control for validating Alomone Labs Anti-ASIC3 Antibody (#ASC-018).

    Overexpressed Membrane Fractions are:
    ✓ Lyophilized powder
    ✓ Economical
    ✓ Shipped at room temperature (no need for dry ice and extra shipping costs)
    ✓ User-friendly & time-saving. Just add water, sample buffer and load your gel

    ASIC3 Channel Overexpressed Membrane Fractions, along with Anti-ASIC3 Antibody, are available as part of the following kit:
    ASIC3 Channel Antibody and Membrane Fractions Kit (#LK-100).

    *Product includes non-injected oocyte membrane lysates

    For research purposes only, not for human use