Facile generation of giant unilamellar vesicles using polyacrylamide gels
Facile generation of giant unilamellar vesicles using polyacrylamide gels
复制标题
使用聚丙烯酰胺凝胶轻松生成巨型单层囊泡
作者：
Eric Parigoris;Daniel L. Dunkelmann;Allan Murphy;N. Wili;A. Kaech;C. Dumrese;Noemi Jiménez;U. Silvan
巨单层囊泡（GUV）是模型细胞大小的系统，其具有广泛的应用，包括药物递送、膜生物物理学分析和细胞机器的合成重建。尽管在过去几十年中已经建立了许多用于产生自由浮动GUV的方法，但只有一小部分成功地从带电脂质和生理离子强度的缓冲液中产生均匀的囊泡群体。在这里描述的方法中，我们通过沉积在软聚丙烯酰胺（PAA）凝胶上的脂质膜的再水化产生大量的自由浮动的GUV。我们表明，这种技术产生高GUV浓度的范围内的脂质类型，包括带电的，独立的缓冲液的离子强度。我们证明，PAA凝胶的温和水合导致主要是单层囊泡，这与本工作中分析的可比方法相反。单层性是GUV的一个定义特征，均匀种群的产生对于许多下游应用来说是关键。PAA方法具有广泛的适用性，并且可以使用常用的实验室试剂轻松实现，使其成为膜生物物理学研究的吸引人的平台。
Giant unilamellar vesicles (GUVs) are model cell-sized systems that have broad applications including drug delivery, analysis of membrane biophysics, and synthetic reconstitution of cellular machineries. Although numerous methods for the generation of free-floating GUVs have been established over the past few decades, only a fraction have successfully produced uniform vesicle populations both from charged lipids and in buffers of physiological ionic strength. In the method described here, we generate large numbers of free-floating GUVs through the rehydration of lipid films deposited on soft polyacrylamide (PAA) gels. We show that this technique produces high GUV concentrations for a range of lipid types, including charged ones, independently of the ionic strength of the buffer used. We demonstrate that the gentle hydration of PAA gels results in predominantly unilamellar vesicles, which is in contrast to comparable methods analyzed in this work. Unilamellarity is a defining feature of GUVs and the generation of uniform populations is key for many downstream applications. The PAA method is widely applicable and can be easily implemented with commonly utilized laboratory reagents, making it an appealing platform for the study of membrane biophysics.
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影响因子：
4.6
作者：
Steinkühler J;De Tillieux P;Knorr RL;Lipowsky R;Dimova R
通讯作者：
Dimova R
影响因子：
15
作者：
Horger, Kim S.;Estes, Daniel J.;Capone, Ricardo;Mayer, Michael
通讯作者：
Mayer, Michael
影响因子：
9.5
作者：
Hu, Peichi C.;Li, Su;Malmstadt, Noah
通讯作者：
Malmstadt, Noah
DOI：
10.1007/978-1-60761-063-2_13
发表时间：
2010
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
--
作者：
Chaudhuri, Tathagata;Rehfeldt, Florian;Sweeney, H Lee;Discher, Dennis E
通讯作者：
Discher, Dennis E
DOI：
10.1073/pnas.96.15.8461
发表时间：
1999-07-20
影响因子：
11.1
作者：
Korlach, J;Schwille, P;Feigenson, GW
通讯作者：
Feigenson, GW