生物医学工程学杂志

生物医学工程学杂志

支撑脂双层膜的制备及其功能化研究

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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 80∶20,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 vitrobecause 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)] (DGS-NTA) 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

Key words: supported lipid bilayers; DOPC and DGS-NTA; fluorescence recovery after photobleaching; diffusion coefficient; fluidity

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