Objective To prepare 18F-5-fluoro-N-(2-(diethylamino) ethyl) picolinamide (18F-5-FPN) and evaluate its binding affinity with melanin.Methods 5-bromo-N-(2-(diethylamino) ethyl)picolinamide (precursor) was synthesized and the structure was characterized by ultraviolet (UV),nuclear magnetic resonance (NMR) and mass spectrometry.18F-5-FPN was prepared with FX-XN module through nucleophilic substitution reaction.The product was purified and identified by HPLC.The binding specificity of 18F-5-FPN with melanin was demonstrated by in vitro study of cellular uptake,and in vivo static PET imaging of pigmented B16F10 and amelanotic A375m allografts.Biodistribution study was performed to evaluate the pharmacokinetics of 18F-5-FPN in vivo.Two-sample t test was used for data analysis.Results The structure of precursor was characterized by UV,1H NMR,13C NMR and mass spectrometry.18 F-5-FPN was successfully prepared with radiochemical yield of 5％-8％,radiochemical purity ＞95％ and the specific activity of 100-120 GBq/μmol.The cell uptake study showed that the uptakes of 18F-5-FPN in B16F10 cells at 30,60 and 120 min ((9.80±0.46) ％,(10.34±0.32) ％,(7.27±0.26)％) were significantly higher than those in A375m cells ((1.36±0.14)％,(1.75±0.12)％,(1.54±0.09)％;t =30.3,46.8,38.4,all P＜0.05),and the optimal uptakes were observed at 60 min for both cells.In static PET imaging,the tumors in B16F10-bearing mice were clearly visible with the uptake value of (18.20±3.21) ％ID/g and the T/B ratio of 19.17±10.03 at 1 h postinjection,while no tumor uptake was seen in A375m-bearing mice.The main clearance pathway of 18F-5-FPN was the renal system,which cleared the unbound tracer rapidly.Conclusions 18F-5-FPN can specifically target the melanin in vitro and in vivo with favorable pharmacokinetics and good T/B ratio.18F-5-FPN may be an ideal molecular probe for diagnosis of malignant melanoma.

Mammalian zona pellucida 3(ZP3) plays an important role in the induction of capacitating sperm acrosome reaction. In this study, we obtained the soluble mZP3 fusion protein and identified its immunoreactivity. mZP3 cDNA was cloned into plasmid pMAL-p2x, and the recombinant plasmid was transformed into Escherichia coli BL21. To get the soluble mZP3 fusion protein, we tried to optimize the expression conditions, including additives, IPTG concentrations, temperatures and induction duration. Then, Western blotting and ELISA were used to identify the immunoreactivity of the purified protein. Based on the optimization experiments, we concluded that the best soluble expression conditions for the mZP3 fusion protein involved incubation to an A600 of 0.6, addition of glucose to a final concentration of 0.02 mol/L, addition of IPTG to a final concentration of 0.6 mmol/L and then further incubation for 4 h at 25 degrees C. Western blotting and ELISA showed that the mZP3 fusion protein retained immunoreactivity. The fusion protein can be used as solubility antigens for developing the immunocontraception vaccines of mZP3 and detecting the immune effects of the vaccine.
Animals ; Egg Proteins ; biosynthesis ; genetics ; immunology ; Escherichia coli ; genetics ; metabolism ; Female ; Membrane Glycoproteins ; biosynthesis ; genetics ; immunology ; Mice ; Receptors, Cell Surface ; biosynthesis ; genetics ; immunology ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Solubility ; Vaccines, Contraceptive ; immunology ; Zona Pellucida Glycoproteins