Rabbit anti syn

As previously described by us,
⁴⁰ (link) proteins were extracted from the MCA‐supplied brain regions and loaded onto SDS‐polyacrylamide gel for electrophoresis. Gel transfer to a PVDF membrane was performed under 200 V for 1 h. Membranes were blocked with 5% skimmed milk. Membranes were incubated with primary antibodies (1:1000, rabbit anti‐NLRP3, Merck; 1:500, rabbit anti‐ASC, Abcam; 1:1000, rabbit anti‐IL‐1β, Abcam; 1:1000, rabbit anti‐IL‐18, Proteintech; 1:1000, rabbit anti‐caspase‐1, Abcam; 1:500, rabbit anti‐GSDMD, Affinity; 1:2000, rabbit anti‐G3BP1, Proteintech; 1:500, rabbit anti‐TIA1, Proteintech; 1:1000, rabbit anti‐DDX3X, Proteintech; 1:1000, rabbit anti‐IgG antibody, Cell Signaling Technology; 1:10,000, rabbit anti‐SYN, Abcam; 1:1000, rabbit anti‐PSD‐95, Cell Signaling Technology; 1:1000, rabbit anti‐BDNF, Abcam; 1:500, rabbit anti‐Ang‐1, Proteintech; 1:500, rabbit anti‐Ang‐2, Proteintech; 1:500, rabbit anti‐VEGF, Abcam; 1:1000, mouse β‐actin, Santa Cruz) for 24 h at 4°C. Next, membranes were incubated with a secondary antibody (1:4000, goat anti‐rabbit IgG, goat anti‐mouse IgG, Cell Signaling Technology) for 2 h at room temperature. Western blot images for each of the antibodies were analyzed using an image analysis program (ImageJ 1.42, National Institutes of Health) to quantify protein expression in terms of relative image density.

At 3, 14, and 28 days after initiation of the exercise regimens, rats were sacrificed for Western blot analysis. Tissue samples from the ipsilesional ischemic cerebral hemispheres of all experimental groups were harvested, and total protein extraction was performed using cell lysis solutions (Thermo Fisher Scientific, Inc., Waltham, MA, USA). Protein concentration was then determined by the BCA method. Electrophoresis (10% SDS-PAGE gel) was performed with 30 μg of protein per lane. Gel transfer to a PVDF membrane was performed under 200 V for 1 h. Membranes were blocked with 5% skimmed milk, followed by incubation with primary antibodies (1:1,000 rabbit anti-BDNF, rabbit anti-NGF, rabbit anti-PSD-95, rabbit anti-SYN, rabbit anti-Tau, and rabbit anti-GAP43, Abcam, MA, USA; 1:500 rabbit anti-HIF-1α, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) overnight at 4°C. The next day, membranes were washed three times and further incubated with a goat anti-rabbit IgG-HRP secondary antibody (1:1,000, Santa Cruz) at room temperature for 1 h. After washing, the ECL method was used to detect signals. Western blot images for each antibody were analyzed using an image analysis program (ImageJ 1.42, National Institutes of Health, Bethesda, MD, USA) to quantify protein expression according to relative image density.