Cleaved caspase 3 asp 175 rabbit polyclonal antibody

Brain and tumor samples were collected at 4 h post-treatment, soaked in RNA later, and conserved at 5 °C. Proteins from tumors and cerebella at P5 were extracted, normalized, separated, and immunoblotted as described [41 (link)]. To evaluate apoptosis, cleaved-caspase-3 (Asp 175) rabbit polyclonal antibody was used (dilution 1:1000 overnight; Cell Signaling Technology, Danvers, MA, USA). Monoclonal antibody against HSP70 (dilution 1:10.000 30 min, Sigma-Aldrich H5147, St. Louis, MI, USA) was used as loading control. Specific proteins were visualized using ECL™ Prime Western Blotting Detection Reagent (Cytiva RPN2232) with ChemiDoc system XRS+ Biorad and quantified using ImageJ software version 1.8.0.

For immunoblot, total proteins were extracted from cerebella (n = 2) at P3. Experimental groups included sham-injected (PBS) mice; mice injected with 0 Gy-plasma-derived exosomes (Evs SI (0 Gy); mice injected with 2 Gy PBI-plasma-derived exosomes (Evs PBI (2 Gy); mice injected with 2 Gy WBI-plasma-derived exosomes (Evs WBI (2 Gy); 2 Gy whole-body-irradiated mice (2 Gy WBI). All brain samples were collected at 6 h post-treatment. Proteins were extracted, normalized, separated, and immunoblotted as described [76 (link)]. To evaluate apoptosis, samples were incubated with Cleaved-caspase-3 (Asp 175) rabbit polyclonal antibody (dilution 1:1000; Cell Signaling Technology, Danvers, MA, USA). Monoclonal antibody against β-actin (dilution 1:10,000, Sigma-Aldrich, St. Louis, MO, USA) was used as a loading control. Specific proteins were visualized with ChemiDoc system XRS+ Biorad and quantified using ImageJ software.