Gapdh rabbit polyclonal antibody

Cells (1.5×10³) from each cell line were lysed in RIPA buffer (Cell Signaling Technology) with 0.5% SDS and complete protease inhibitors (Roche) by mechanical disruption and freeze thawing; the cell mixtures were then reduced and denatured in NuPage LDS Sample Buffer (Life Technologies) with 8% β-mercaptoethanol, at 95°C for 5 minutes. The samples were run next to the Precision Plus Duel Color Standards (Bio-Rad) in a 4–12% Bis-Tris polyacrylamide gel (Life Technologies). Expression of each reporter protein and self-cleavage of the viral 2A sequences were evaluated by Western blots with a DsRed rabbit polyclonal antibody (Clonetech) diluted 1∶3000, firefly luciferase mouse monoclonal antibody (Abcam) diluted 1∶3000, HSV-1 thymidine kinase goat polyclonal antibody (Santa Cruz Biotechnology) diluted 1∶250, and GAPDH rabbit polyclonal antibody (Sigma-Aldrich) diluted 1∶5000. Secondary antibodies used were a goat anti-rabbit IgG HRP conjugate diluted 1∶3000 (Cell Signaling Technologies), goat anti-mouse IgG HRP conjugate diluted 1∶3000 (Bio-Rad), and donkey anti-goat IgG HRP conjugate diluted 1∶2500 (Promega).

Sperm were lysed with radioimmunoprecipitation assay buffer containing protease and phosphatase inhibitors (Thermo Fisher Scientific). The polyvinylidene difluoride membranes were incubated with specific antibodies including CISY rabbit mAb (Cell Signaling Technology, 14309S), FUMH rabbit polyclonal antibody (Proteintech, 11375-1-AP), ODO1 rabbit polyclonal antibody (Proteintech, 15212-1-AP), SDHA rabbit mAb (Cell Signaling Technology, 11998T), CPT2 rabbit mAb (Abcam, ab231162), ATPA mouse mAb (Invitrogen, 459240), GAPDH rabbit polyclonal antibody (Sigma-Aldrich, G9545), PPARγ rabbit polyclonal antibody (Invitrogen, PA3-821A). Protein bands were measured using an Odyssey Infrared Imaging System (ODYSSEY CLx, Li-COR). The fluorescence intensity was evaluated using Image Studio Lite version 5.2 (https://www.licor.com/bio/image-studio-lite/).