Objective:

A protein to aequorin (the chemiluminescent protein from Aequorea named jellyfish) green fluorescent protein (GFP) has become established as an important tool in drug discovery and biological research (1). It consists of 238 amino acids and its molecular mass is 27-30 kDa. GFP fluorescence occurs without cofactors and this property allows GFP fluorescence to be utilized in nonnative organisms. Genetically engineered cells with GFP expression have provided a valuable tool for automated analysis, and can be adapted for high-throughput systems (2). Inorganic NPs, including metal oxides, are promising materials for applications in medicine, such as cell imaging, biosensing, drug/gene delivery, and cancer therapy. Zinc oxide (ZnO) NPs belonging to a group of metal oxides are characterized by their photocatalytic and photo-oxidizing ability against chemical and biological species. In recent times, ZnO NPs have received much attention for their implications in cancer therapy (4). ZnO nanoparticles have even been shown to specifically target cancer cells and can possibly be developed as an alternative anticancer therapeutic agent. Although several studies have already characterized the toxicology of ZnO nanoparticles in vitro with a variety of mammalian cell lines, most of these studies focused on the cytotoxicity of ZnO nanoparticles as a means of assessing biocompatibility (5). ZnO utilizes a multifunctional nanoplatform that bombards malignant cells from the outside. On the other hand, ZnO nanoparticles propose new opportunities including the improvement of the specific drug delivery and also manipulating cell membranes. No cytotoxic effects of ZnO nanoparticles were found in human glioma cells. ZnO nanoparticles are known to be one of the multifunctional inorganic nanoparticles with effective antibacterial activity (6).

Methods:

In this study, ZnO/GFP was synthesized previously, used in cell imaging studies for targeting and labeling platform. In the synthesis procedure, initially ZnO/GFP was conjugated through crosslinker. The bonding of GFP to the ZnO nanoparticles yield was detected by High Performance Liquid Radio Chromatography. The complex was labeled with 131I via iodogen method. The yield of radiolabeling of ZnO/GFP was determined by Thin Layer Radio Chromatography. 131I labeled ZnO/GFP (131I/ZnO/GFP) was added to the glioma cells in order to investigate its optical signals. Cytotoxicity studies were carried out on Human glioblastoma cells (U87-MG), 24 hour by MTT method.

Results:

The sizes of ZnO were change by the incubation time. Radiolabeling yield of 131I/ZnO/GFP was 98.42±0.90%. There are no toxicity till to concentration 1000 ng/ml limit. U87-MG were used to determinate the time dependent incorporation of 131I, 131I/ZnO and 131I/ZnO/GFP. Due to investigation time dependent incorporation of 131I 0.037 MBq (1.0 µCi/mL) labeled components, the cells were incubated for 1, 2, 4 and 24 hour at 37 °C. Two hour was found as the maximum uptake time.

Conclusion:

All experimental results suggest that 131I labeled GFP/ZnO compound can be used in the applications of Nuclear Medicine as a radiolabeled agent for imaging both optical and radioisotopical.

Keywords: Green Fluorescent Proteins (GFP),zinc oxide nanoparticles,131I,glioma cells (U87-MG)