Egfr-targeted Ultrasmall Superparamagnetic Iron Oxide (uspio) Particle For Magnetic Resonance Molecular Imaging Of Lung Cancer Cell In Vitro

ObjectivePreparation of the molecular imaging agent C225-USPIO targeting EGFR;to analyse targeted effect of contrast agent on A549 cells and proper content.Methods1. Attach ultra small superparamagnetic iron oxide particles USPIO with Cetuximab C225, anti-human IgG to prepare molecular MR contrast agent C225-USPIO and IgG-USPIO through Carbodiimide method, the latter as the control group, and set different concentrations according to Fe content;2. Culture lung cancer A549 cells, immunocytochemistry (SP) to detect the expression of EGFR on cells;3. Detect the binding rate of A549 cells with C225-USPIO by immunofluorescence staining and flow cytometry, measure antibody activity of the contrast agent, and to explore the best contrast about Fe concentration;4. According to Fe concentration of 50μg/ml, culture A549 cell and C225-USPIO, detect the activity of the circumstances about C225-USPIO into the cells after 1 hour with Prussian Blue staining and TEM ; 5. Detect imaging effects of contrast agent targeting on cells with 4.7T MRI, scanning were marked as the contrast agent C225-USPIO cells, IgG-USPIO cells, and distilled water, scanning sequences included axial T1WI, T2WIResults1. Immunocytochemical detection of lung cancer A549 cells is positive for EGFR expression;2. Immunofluorescence staining and flow cytometry showed the binding rate higher than the control group after cells cultivation with different concentrations of C225-USPIO, when the Fe concentration of 0.1mg/ml, the fluorescence intensity and the binding was the best;3. Prussian Blue staining and Transmission electron microscopy revealed that in C225-USPIO contrast agent group of cells the particle content of Fe in cytoplasmic vesicles or on surface was more than the control group;4. 4.7T MRI scan revealed C225-USPIO group of cells decreased significantly more than unlabeled cells about the T2WI signal, but there was no significant difference between the time gradient.Conclusion We construct the molecular imaging agent C225-USPIO targeting on EGFR of A549 lung cancer cells successfully , the imaging agent showed good targeting effect and specificity, and reduced MRI T2 value significantly, such molecular contrast agent could provide a new way measurement of EGFR expression level, also provide the basis for the next step nude mice experiments.