Affibody靶向载自杀基因脂质纳米超声造影剂的制备及其特性的实验研究

Preparation and experimental study of Affibody targeting suicide gene carrying nano ultrasound contrast agents

目的:制备一种包裹自杀基因和液态氟碳并具有Affibody靶向以及核定位效应的脂质纳米超声造影剂(UCAs),研究其基本特性及超声显影效果。方法:以前期课题组构建的含二硫键的聚氨基胺类聚合物CHL-CBA为核定位载体,构建基因载体复合物,采用薄膜水化机械振荡法制备包裹该复合物和液态氟碳的脂质纳米UCAs,通过生物素-亲和素法将Affibody连于脂质纳米UCAs表面,激光共聚焦显微镜检测Affibody与纳米UCAs连接情况,并对其外观形态、粒径、电位等进行表征。观察靶向纳米UCAs体外超声显影效果。结果:Affibody靶向载自杀基因和液态氟碳的纳米UCAs平均粒径为(426.8±66.45)nm,Zeta电位为(-11.5±3.04)m V,大小均一,呈球形,Affibody与其连接成功。体外超声显像显示出较好的超声对比增强能力,呈均匀细腻的密集高回声。结论:本实验成功制备了Affibody靶向载自杀基因脂质纳米UCAs,体外显影效果良好,为后续体内肿瘤靶向超声造影奠定了基础。

Objective: To prepare a kind of lipid nano ultrasonic contrast agents(UCAs) with Affibody targeting and nuclear localization effect wrapped suicide gene and liquid fluorocarbon, and to study its basic characteristics and ultrasonic development effect. Method: To check and ratify the carrier using the previous team construction of polyamino amine polymers containing disulfide bond(CHL-CBA), to build complex gene carrier using thin-film hydration mechanical oscillation of the packages of fluorocarbon compounds and liquid lipid nanoparticle UCAs, to connect Affibody to lipid nanoparticle UCAs surface using biotin avidin method, to detect the connection of Affibody and nano UCAs using laser confocal microscopy, and to characterize the appearance of the morphology, particle size, potential, etc, and to observe the effect of ultrasonic development of targeted nanometer UCAs in vitro. Results: The average particle diameter of UCAs was(426.8±66.45)nm, and the Zeta potential was(-11.5±3.04)mV. The UCAs were spherical in size, which was successfully connected with Affibody. In vitro, it showed a good contrast-enhanced ability of ultrasound, presenting uniform and fine dense hyperecho. Conclusion: In this study, the Affibody targeting suicide gene carrying lipid nanoparticles UCAs were successfully prepared, and the in vitro development effect was good, which laid a foundation for the follow-up in vivo tumor targeted contrast-enhanced ultrasound.