γ h2aχ mouse monoclonal primary antibody

The CT26 tumor-bearing mice with tumor volume of 150–200 mm³ were classified: Vehicle, GGd-NCPs, aAGd-NWs ([Gd] = 15.7 mg kg⁻¹, [ara-AMP] = 34.7 mg kg⁻¹) with or without X-ray irradiation (5 Gy × 1). The corresponding drugs were administered intravenously 6 h before radiotherapy. Then, tumor tissues were collected for evaluation of γ-H2Aχ 24 h post irradiation. Tumor tissues were sliced and stained with γ-H2Aχ mouse monoclonal primary antibody (diluted 1:200 with 3% BSA, Abcam, UK) and secondary antibody conjugated to Alexa Fluor 488 (Bioss, China) to detect DNA double-strand breaks. According to the manufacturer’s protocol, tumors were collected from six groups at 48 h post different treatments for terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay and Ki67 staining, respectively.

CT26 cells were seeded in confocal dishes at 2 × 10⁵ per dish and incubated overnight for attachment, which were divided into six groups of Vehicle, Vehicle+RT, Gd-NCPs, GGd-NCPs+RT, aAGd-NWs, aAGd-NWs+RT ([Gd] = 50 μM, [ara-AMP] = 50 μM). After incubation for 6 h, the cells were treated with radiation (0 Gy or 5 Gy × 1). Two hours after irradiation, all treatments were removed, and tumor cells were fixed with 4% paraformaldehyde and washed with PBS. Then, 0.3% Triton-X was used to perforate the nucleus. Then, cells were added with 1% bovine serum albumin solution as a blocking buffer, and incubated with γ-H2Aχ mouse monoclonal primary antibody (1:500, Abcam, UK) for 1 h and washed with PBS. After that, Alexa Fluor 488 conjugated secondary antibody (1:500, Bioss, China) was added and incubated for 1 h. Then, DAPI was used to stain the nucleus. Tumor cells were observed under Olympus FV3000 CLSM, and analyzed by ImageJ software.