For 64Cu-diacetyl-bis(N4-methylsemicarbazone) (64Cu-ATSM) high-specific-activity 64Cu radioisotope was obtained from Washington University School of Medicine, and ligand synthesis and labeling was conducted as previously described (9 (link)). A radiochemical purity of 99% was assessed by instant thin-layer chromatography. 18F-fluoride was produced on the MSKCC cyclotron (TR19/9; EBCO Technologies, Inc.). 18F-fluoromisonidazole (18F-FMISO) (specific activity, 370 MBq/µg [10 mCi/µg]) was prepared according to the method of Grierson et al. (10 (link)) by the Cyclotron and Radiochemistry Service at MSKCC. 18F-fluoroazomycin arabinoside (18F-FAZA) was prepared as described according to the method of Maier et al. (11 (link)). The radiochemical purity of 18F-FMISO and 18F-FAZA, determined by high-performance liquid chromatography, was greater than 98%. 18F-flortanidazole (18F-HX4) was obtained from Siemens PETNET solutions (Siemens Medical Solutions, USA) and radiochemical purity greater than 98% by high-performance liquid chromatography.
Animals were intravenously coadministered 9.25–33.3 MBq (250–900 µCi) of the respective tracer along with pimonidazole hydrochloride (80 mg/kg; Hypoxyprobe-1 [Natural Pharmacia International Inc.]), in a final injection volume of 200 µL. Hoechst 33342 trihydrochloride (Sigma; 1 mg in 100 µL of physiologic saline) was injected 1 min before sacrifice.
Animals were intravenously coadministered 9.25–33.3 MBq (250–900 µCi) of the respective tracer along with pimonidazole hydrochloride (80 mg/kg; Hypoxyprobe-1 [Natural Pharmacia International Inc.]), in a final injection volume of 200 µL. Hoechst 33342 trihydrochloride (Sigma; 1 mg in 100 µL of physiologic saline) was injected 1 min before sacrifice.