Figure 3
Radiolabeling and quality control of 131I-ANAs. (a )Efficiency yield of 131I-ANAs as determined by paper electrophoresis in 0.025 M phosphate buffer and radioautography (Cyclone, PerkinElmer). Chromatograms were analyzed using OptiQuant 5 software. 131I-ANAs stayed at the anode Rf = 0.0–0.25, and free 131I was separated from the reaction mixtures and migrated to the cathode Rf = 0.75–0.85. (b) 131I-ANAs were collected in fraction no. 3–6 mL after purification through gel sephadex, and free 131I was collected in fraction no. 8–10 mL. (c) Radiochemical purity of 131I-ANAs as determined by Tec-Control-Chromatography (TCC 150–771), Biodex. The strip was placed into a vial containing 0.9% saline, and the strip was developed until the solvent front migrated to the solvent front line. The strip was removed, and then the storage phosphor screen was used. Chromatograms were analyzed using OptiQuant 5 software. 131I-ANAs remained at the origin Rf = 0.0–0.3, and free 131I was separated from the reaction mixtures and migrated to the front Rf = 0.9–1.0. (d) Hypothetical chemical structures of 131I-ANA conjugates,131I attachment to the ortho site of the phenol ring tyrosine. (e) Stability of 131I-ANAs in 0.9% NaCl and 0.05 M PBS after cryopreservation and storage 4 °C. All experiments were performed in duplicate and 3 independent times (n = 3). Simple linear regression analysis of the results was performed by GraphPad Prism Software 8. (f) 131I-ANAs in 0.05% human serum were stable (96.3 ± 0.31% radiochemical purity) after incubation at 37 °C.
