AnnexinV纳米级超声造影剂的制备及体外靶向显像的实验研究  

Study of preparation of the Annexin V-nanoscale ultrasound contrast agents and targeting ultrasound imaging in vitro

作　　者：周田[1] 赵萍[1] 段云友[1] 蔡文斌[1] 杨恒丽[1] 张惠中[2] 刘冲[2] 

机构地区：[1]第四军医大学唐都医院超声诊断科,西安710038 [2]第四军医大学唐都医院临床实验与检验科,西安710038

出　　处：《中华超声影像学杂志》2015年第12期1064-1070,共7页Chinese Journal of Ultrasonography

基　　金：基金项目：国家自然科学基金（81201136）

摘　　要：目的探讨连接AnnexinV纳米级超声造影剂的制备、超声显影及体外与肿瘤凋亡细胞的结合能力。方法采用薄膜水化法制备包裹医用全氟丙烷气体的纳米级脂质微泡，通过生物素一亲和素系统连接AnnexinV分子，制成AnnexinV纳米超声造影剂（AnnexinV—Nanobubbles，AVNBs）；纳米粒径／电位分析仪检测AVNBs的粒径和Zeta电位；将AVNBs分别于4℃存放0h、12h、24h、72h后测粒径，并分析其稳定性；扫描电子显微镜观察其形态及均一度；超声成像系统检测其体外显影效果，并与声诺维进行对比；荧光显微镜观察AVNBs与体外肿瘤凋亡细胞的结合情况。结果测得AVNBs粒径大小为（640．2±32．1）nm，Zeta电位为（-23．30±5．71）mV，且24h内粒径基本稳定在纳米级；肉眼观察制备好的AVNBs为乳白色混悬液。扫描电镜视野下呈空心球形、大小均一、分散良好。体外超声成像显示，AVNBs与对照组声诺维有相同的显影效果。体外实验证实AVNBs与凋亡细胞结合良好，结合率为（97．55±1．30）％。结论本法制备的AVNBs粒径小，稳定性和均一性好，体外超声显影效果明显，可与体外凋亡细胞靶向性结合。Objective To research the Annexin V-nanoscale ultrasound contrast agents＇ preparation, ultrasound imaging and the ability to binding apoptosis cells of tumor in vitro. Methods The nanoscale bubble （Nanobubbles, NBs） packaged the octaflouropropane （C3F8） gas was prepared by thin film hydration. The Annexin V-Nanobubbles （AVNBs） solutions was acquired through conjugating the biotinylated-Annexin V to the surface of the NBs by biotin-streptavidin bridging chemistry. The size and zeta potential of AVNBs were measured by NanoPlus-3 zeta/nano particle analyzer. The shift in size distribution of AVNBs bubbles was analyzed for the stability,after it was stored at 4 ℃ for different time. AVNBCs shape were measured by scanning electron microscopy. The AVNBs bubble was measured using an ultrasound system for echogenicity in vitro, and SonoVue was for control. Finally, the ability of AVNBs binding with apoptosis cells of tumor in vitro was determine via the fluorescence microscope. Results AVNBs has a size distribution of （640.2 ± 32.1）nm,and a mean zeta potential of （ - 23.30 ±5.71） mV. Its size remained relatively constant and appeared to show less size variation within the 24 h analysis period. AVNBs solutions were visible milky white and slightly suspension liquid with the naked eye. Under scanning electron microscopy, AVNBs were uniform hollow sperical cavitation bubble with small size and larger dispersibility in solution. The AVNBs and SonoVue solution had the same higher grayscale signal intensity by ultrasonic imaging. The AVNBs binded well with apoptosis ceils of tumor in vitro,and the rate of binding was （97.55 ± 1.30）%. Conclusions The AVNBs particles prepared by method of thin film hydration have a nanoscale size,good stability and echogenicity. It can be targeted binding with the apoptosis cells of tumor in vitro.

关 键 词：微气泡 纳米技术 细胞凋亡 脂联蛋白A5 

分 类 号：R943[医药卫生—药剂学] R96[医药卫生—药学]