Font Size: a A A

Preparation And In Vitro And In Vivo Experimental Study Of STEAP-1-targeted Ultrasound Contrast Agent

Posted on:2017-09-02Degree:MasterType:Thesis
Country:ChinaCandidate:Y LiuFull Text:PDF
GTID:2334330485476396Subject:Medical imaging and nuclear medicine
Abstract/Summary:PDF Full Text Request
Objective: To prepare the six-transmembrane epithelial antigen of the prostate(STEAP-1)targeted novel ultrasound contrast agent via biotin-streptavidin binding and to evaluate the binding rate,stability and searching target function in vitro.The specific contrast enhancement using the STEAP-1 targeted novel microbubbles in the two kinds of tumor-bearing nude mice was analyzed to identify the targeted function in vivo.Methods:(1)The LNCa P and PC3 were cultured to detect the expression of STEAP-1 by immunofluorenscenece assay and western blot.(2)The human STEAP-1 antibody was conjugated to the Sono Vue microbubbles using avidin-biotin complex technology and then physiochemical properties of the targeted microbubbles were observed using optical and fluorescence microscope.The binding rate and stability of microbubbles were examined using flow cytometry.(3)To detect the bining capacity of the prepared targeted microbubbles in vitro after the full reaction with the slides of prostate cancer cells by immunofluorenscenece assay.(4)To monitor the contrast enhancement in real time by an ultralsound system in LNCa P and PC3 transplanted tumors in nude mice.To compare and analyze the peak intensity(PI),time to peak(TTP),area under the curve(AUC)and mean transit time(MTT)of targeted and blank nanobubbles.Results:(1)Under fluorescence microscope the membranes of prostate cancer cells,LNCa P and PC3,exhibited green fluorescence.The western blot assay revealed that the expression of STEAP-1 in LNCa P is higher than in PC3.(2)Under fluorescence microscope the surface of STEAP-1 targeted microbubbles seemed like green circles,while no green fluorescence was observed on the surface of the control microbubbles.and The prepared targeted microbubbles had a same bubble size and distributed evenly under the microscope.The flow cytometry showed the targeted one had a higher binding rate than the control one,and the difference was statistically significant(p<0.05).And the STEAP-1-carrying rate had no obvious change before and after shaking in vitro,but the difference was not statistically significant(p>0.05).(3)The targeted microbubbles showed a concentration in the cell regions after the reaction with the prostate cancer cells.After washing the slides with PBS buffer,a large amount of the targeted microbubbles adhered to the surface of LNCa P and PC3,which seemed like garlands.No adhesion of control microbubble was observed around the cells.The LNCa P had a higher adherence rate in comparison with PC3,the difference was statistically significant(p<0.05).And their rates had no significant change after washing again(p>0.05).(4)The contrast enhancement imaging showed echo enhancement evidently and then echo waning gradually in the transplanted tumor tissues.There was significant difference between the targeted and control microbubbles in the PI,AUC and MTT(p<0.05)in two transplanted prostate tumors,but the TTP was not significantly different(p>0.05).Among the two kinds of transplanted tumors with targeted microbubbles,the PI and AUC in LNCa P tumors was significantly higher than that in PC3 tumors(p<0.05).Conclusion:(1)The expression of STEAP-1 is both positive in LNCa P and PC3,and the representative cell line of early prostate cancer,LNCa P,shows a higher expression.(2)The STEAP-1 targeted novel microbubbles can be prepared successfully via avidin-biotin complex technology with a high binding rate and a good stability.(3)In vitro the STEAP-1 targeted novel microbubble is combined with the prostate cancer cells firmly,and the LNCa P also has a higher binding rate compared with PC3.(4)The STEAP-1 targeted microbubble has a high specificity in vivo,which illustrates that the STEAP-1 can be a new ideal target for ultrasound targeted contrast imaging and it has special value for basic research and clinical application of prostate cancer.
Keywords/Search Tags:STEAP-1, targeted microbubbles, prostate carcinoma, biotin-avidin, contrast-enhanced ultrasound
PDF Full Text Request
Related items