支撑脂双层膜的制备及其功能化研究  

The preparation of supported lipid bilayers and its functional study

作　　者：邹卉 吴建华[1] 方颖[1] ZOU Hui;WU Jianhua;FANG Ying(School of Bioscience &Bioengineering,South China University of Technology,Guangzhou 510006,P.R.China)

机构地区：[1]华南理工大学生物科学与工程学院,广州510006

出　　处：《生物医学工程学杂志》2019年第1期85-93,共9页Journal of Biomedical Engineering

基　　金：国家自然科学基金(11672109;11432006;11272125);中央高校专项基金(华南理工大学D2156960)

摘　　要：支撑脂双层膜(SLBs)因其结构与功能均类似天然细胞膜而广泛应用于生物医学及生物工程研究。本文以玻璃载玻片为基底,以1, 2-油-锡-磷脂-3-卵磷脂(DOPC)与{亚氨基二乙酸丁二酰(镍盐)}(DGS-NTA)为原料,采用荧光漂白恢复技术(FRAP)研究原料配比、制备方法、漂白区域大小及蛋白加载浓度对SLBs流动性及扩散系数的影响。结果表明:①声波破碎法优于薄膜挤压法,预处理脂质体得到的小囊泡粒径更小、更均匀,并且SLBs形成过程中须避免接触空气;②膜的荧光恢复速度和扩散系数随漂白区域的增大而减小,随着DOPC与DGS-NTA摩尔质量比由98∶2变为84∶16,SLBs流动性及荧光恢复率逐渐减小,当两者配比为82∶18时,膜已失去流动性;③加载蛋白浓度在10～40 nmol·L^(–1)范围时,SLBs的荧光强度随蛋白浓度线性增加,蛋白在SLBs上具有很好的流动性。该研究为细胞膜分子之间的相互作用及后续信号响应的研究搭建了一个良好的生物膜平台。Supported lipid bilayers(SLBs) have been widely used in biomedical and bioengineering research in vitro because its structure and function are similar to natural cell membrane.A fluorescence recovery after photobleaching(FRAP) technique was used to measure the lateral diffusion of the SLBs composed of 1,2-dioleoyl-sn-glycero-3-phosphocholine(DOPC) and 1,2-dioleoyl-sn-glycero-3-[(N-(5-amino-1-carboxyp-entyl) iminodiacetic acid)](DGSNTA) on the glass slide,and the effects of the DOPC-to-DGS-NTA ratio,small unilamellar vesicles(SUV) producing method,sizes of bleaching areas and concentrations of loading proteins on the SLBs fluidity and diffusion coefficient were studied systematically in this paper.The results demonstrated that:(1) SUV made by probe sonication exhibited more uniform and smaller size compared with that made by film extrusion,but the whole process of SLBs formation must not be exposed to air.(2) The fluorescence recovery rate and diffusion coefficient of the SLBs decreased with the increasing bleaching area size.With the mole ratio of DOPC to DGS-NTA decreasing from 98∶2 to 84∶16,the fluidity and fluorescence recovery degree decreased gradually,and the SLBs would lose its fluidity if the ratio reached to 82∶18.(3)The average fluorescence intensity of SLBs increased linearly with the loading protein concentration(10–40 nmol·L–1),and the protein showed good mobility on the SLBs.The study would provide a good platform of bio-membrane for further research on interactions among cell membrane molecules and subsequent signals response.

关 键 词：支撑脂双层膜 DOPC和DGS-NTA 荧光漂白恢复 扩散系数 流动性能 

分 类 号：R318.08[医药卫生—生物医学工程]