A Method for High-Throughput Measurements of Viscosity in Sub-micrometer-Sized Membrane Systems
A Method for High-Throughput Measurements of Viscosity in Sub-micrometer-Sized Membrane Systems
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DOI：
10.1002/cbic.201900510
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发表时间：
2019-12-02
期刊：
影响因子：
3.2
通讯作者：
Saenz, James Peter
中科院分区：
文献类型：
--
作者：
Chwastek, Grzegorz;Petrov, Eugene P.;Saenz, James Peter
为了揭示细菌细胞等小系统中膜适应的基本原理，需要强有力的方法来表征膜流动性。目前可用的相关方法需要先进的仪器，并不适合高通量的设置，需要阐明的生物化学途径参与适应。我们开发了一种快速，稳健，经济实惠的定量方法，使用市售的酶标仪测量散装悬浮液中脂质膜的微粘度。我们的方法，这是适合于高通量筛选，是基于同时测量的吸光度和荧光发射的粘度敏感的荧光染料，9-（2，2-二氰基乙烯基）久洛利定（DCVJ），纳入脂质膜。我们验证了我们的方法，使用人工膜与各种脂质组合物在一系列的温度和观察值与先前发表的结果吻合良好。使用我们的方法，我们能够检测到反刍动物病原体丝状支原体的脂质相变。
To unravel the underlying principles of membrane adaptation in small systems like bacterial cells, robust approaches to characterize membrane fluidity are needed. Currently available relevant methods require advanced instrumentation and are not suitable for high-throughput settings needed to elucidate the biochemical pathways involved in adaptation. We developed a fast, robust, and financially accessible quantitative method to measure the microviscosity of lipid membranes in bulk suspension using a commercially available plate reader. Our approach, which is suitable for high-throughput screening, is based on the simultaneous measurements of absorbance and fluorescence emission of a viscosity-sensitive fluorescent dye, 9-(2,2-dicyanovinyl)julolidine (DCVJ), incorporated into a lipid membrane. We validated our method using artificial membranes with various lipid compositions over a range of temperatures and observed values that were in good agreement with previously published results. Using our approach, we were able to detect a lipid phase transition in the ruminant pathogen Mycoplasma mycoides.