Rabbit monoclonal anti grp78 antibody

The expression of the ERS indicators glucose-regulated protein 78 (GRP78), C/EBP homologous protein/growth arrest and DNA damage-inducible gene 153 (CHOP/GADD153), phosphorylated (p)-eukaryotic translation initiation factor 2 (eIF2α), and caspase 12 was measured by immunohistochemistry staining. Paraffin-embedded brain tissue sections (n = 6 per group) were prepared as described previously (see the TTC staining section). Briefly, sections were deparaffinized and hydrated in decreasing concentrations of alcohol and then incubated for 15 min in 1% Triton X-100 to disrupt the cell membrane. The sections were incubated with 3% H₂O₂ and 5% normal goat serum at room temperature for 30 min each. The brain sections were incubated overnight at 4°C with a rabbit monoclonal anti-GRP78 antibody (1 : 300, Cell Signaling Technology, USA), rabbit polyclonal anti-CHOP/GADD153 antibody (diluted 1 : 50; Sigma-Aldrich, USA), rabbit monoclonal anti-p-eIF2α antibody (1 : 100, Cell Signaling Technology, USA), and rabbit polyclonal anti-caspase 12 antibody (diluted 1 : 50; Abcam, UK). The rest of the protocol was performed following standard procedures. The brain sections were photographed (Leica DM4000, Germany), and the IODs of GRP78, CHOP/GADD153, p-eIF2α, and caspase 12 in the ischemic cortex penumbra were semiquantitatively analyzed by using image analysis software (ImageJ, NIH).

Rabbit monoclonal anti-FMNL2 antibody, rabbit monoclonal anti-Arp2 antibody, mouse monoclonal anti-profilin1 antibody were from Santa Cruz (Santa Cruz, CA, USA). Rabbit monoclonal anti-Fascin antibody was purchased form Abcam (Cambridge, UK). Rabbit polyclonal anti-INF2 antibody was purchased from Proteintech (Proteintech, CHI, USA). Rabbit monoclonal anti-α-tubulin (11H10) antibody, rabbit monoclonal anti-Grp78 antibody, rabbit monoclonal anti-cofilin antibody, and rabbit monoclonal anti-Chop antibody were from Cell Signaling Technology (Beverly, MA, USA). Mouse monoclonal anti-α-tubulin-FITC antibody was from Sigma-Aldrich Corp (St. Louis, MO, USA). Fluorescein isothiocyanate (FITC)-conjugated goat anti-rabbit IgG were from Zhongshan Golden Bridge Biotechnology (Beijing). ER-Tracker Red, Mito-Tracker Green, and enhanced mitochondrial membrane potential assay Kit were from Beyotime Biotechnology (Shanghai). All other chemicals and reagents were from Sigma-Aldrich Corp, unless otherwise stated.