SonoVue与脂质体耦联超声微泡复合载体的制备及其效率研究

Construction methods and its efficiency of the delivery vehicle： complex of SonoVue conjugated to liposomes

目的建立商品化超声造影剂SonoVue与自制脂质体耦联的方法，并检测微泡携带脂质体的效率。方法通过在SonoVue脂质膜中嵌入DSPE～PEG（2000）Amine，使微泡壁带有活性氨基，以戊二醛交联法使其与脂质体耦联形成脂质体-微泡复合载体，并检测影响其耦联效率的两个主要因素。结果成功构建该新型复合微泡造影剂，每200μl微泡混悬液内分别加入150μlDSPE-PEG（2000）Amine和200μl戊二醛-脂质体时，耦联效率最高。结论商品化造影剂SonoVue微泡可被修饰，并实现与脂质体耦联。

Objective To build the schema of combination between commercially available ultrasound contrast agent SonoVue and self-made liposomes,and determine its efficiency. Methods Microbubbles was labeled by carbocyanines dye DiI. The effect of DSPE-PEG-FITC labelling was determined by flow cytometer to evaluate the insertion efficiency of the amphipathic molecule DSPE-PEG（2000）Amine into lipid monomers shell. Aminated fluorescent liposomes were prepared by rotary evaporation and their size were determined by laser particle size analyzer. Liposomes and microbubbles were combined by two-step glutaraldehyde crosslinking. The impact of liposome concentration and DSPE-PEG （2000）Amine concentration on the construction efficiency of the complex of SonoVue conjugated to liposomes was determined by flow cytometer and multifunctional ELIASA respectively. Results The configuration of SonoVue microbubbles labeled by DiI and DSPE-PEG-FITC showed fine. The combination between liposomes and microbubbles was realized by glutaraldehyde crosslinking. The flow cytometer showed that 200μl liposomes solution per 200μl microbubbles suspension was the optimal proportion, with the peak positive rate as （87.80 ± 5.9l ）%. Multifunctional ELIASA showed that 150 μl DSPE PEG（2000）Amine （100 μm） solution per 200 μl microbubbles suspension was the optimal proportion, with the peak carry efficiency as （83.41± 2.21）%. Conclusions The amination of commercially available SonoVue is realized by the insertion of DSPE-PEG （2000） Amine molecule. Glutaraldehyde crosslinking the liposomes and microbubbles is feasible. Moreover, we can choose other active groups to modify the microbubbles and liposomes, construct more convenient methods with higher efficiency.