Anti calmodulin

Homogenates of hippocampal tissue were prepared in RIPA buffer containing 50 mM Tris–HCl, pH 7.4, 150 mM NaCl, 2 mM EDTA, 1% sodium deoxycholate, 1% Triton X-00, 1 mM PMSF, 50 mM sodium fluoride, 1 mM sodium vanadate, 1 mM DTT, and protease inhibitors cocktails. PSD fractions were directly dissolved in 1× SDS-PAGE sample buffer. All the protein samples were boiled in 100 °C water bath for 10 min before western blot. The homogenates were resolved on SDS-PAGE and transferred to nitrocellulose membranes, which were incubated in the TBS buffer containing 0.1% Tween-20 and 5% milk for 1 h at room temperature before incubation with a primary antibody overnight at 4 °C. After wash, the membranes were incubated with an HRP-conjugated secondary antibody in the same TBS buffer for 1 h at room temperature. Immunoreactive bands were visualized by ChemiDocTM XRS + Imaging System (BIO-RAD) using enhanced chemiluminescence (Pierce) and analyzed with Image J (NIH). The primary antibodies used were as follows: anti-Histone H3, Cell Signaling (9715); anti-Calmodulin, Millipore (05-173); anti-CaMKIIα, Cell Signaling (11945); anti-p-CaMKIIα Thr²⁸⁶, Sigma (SAB4300228); anti-Acetyllysine, Cell Signaling (9441); anti-p-GluR1 Ser⁸³¹, Abcam (ab109464); anti-GluR1, Abcam (ab109450); anti-GST, Abmart (12G8); and anti-His, Abmart (10E2).