mouse anti-post synaptic density protein 95 (psd-95)  (Novus Biologicals)

Journal: Journal of neurochemistry

Article Title: PI3K modulates estrogen-dependent facilitation of colon-to-urethra cross-organ reflex sensitization in ovariectomized female rats.

doi: 10.1111/j.1471-4159.2010.06577.x

Figure Lengend Snippet: Fig. 2 Effects of intrathecal ICI 182780 and LY294002 pre-treatments on estradiol- dependent up-regulation of phosphorylated Akt (pAkt) and phosphorylated NR2B (pNR2B) in left lumbosacral (L6–S2) dorsal horn tissue of ovariectomized rats. Six hours after subcutaneous estradiol injec- tions (OVXE), expression levels of (a) pAkt and (b) pNR2B both increased when com- pared with vehicle injections (OVXV, **p < 0.01 to OVXV, n = 4). Pre-treatments with ICI 182780 (ICI + OVXE) and LY294002 (LY + OVXE), 30 min before estradiol, reversed the increases of pAkt and pNR2B expression caused by estradiol injections (##p < 0.01 to OVXV, n = 4). (c) Co-immunoprecipitation analysis of left lumbosacral (L6–S2) dorsal horn tissue obtained from OVXV and OVXE animals. Immunoblotting (input) in the left column shows increases in expression levels of pAkt, PSD95, and pNR2B in OVXE when compared to OVXV rats. Immunoprecipita- tion blotting (IP) in the right column shows an increment of PSD95 and pNR2B immu- noprecipitation with anti-pAkt antibody in crude membrane extract from the OVXE but not the OVXV group.

Article Snippet: Binding proteins were eluted with SDS–PAGE sample buffer at 95 C. Proteins were separated by SDS–PAGE, transferred to nitrocellulose membranes electrophoretically, and detected using rabbit polyclonal anti-pAkt, mouse monoclonal anti-post-synaptic density protein (PSD95) (Santa Cruz), and rabbit polyclonal antiNR2B (Millipore).

Techniques: Expressing, Immunoprecipitation, Western Blot, Membrane