STED microscopy detects and quantifies liquid phase separation in lipid membranes using a new far-red emitting fluorescent phosphoglycerolipid analogue
STED microscopy detects and quantifies liquid phase separation in lipid membranes using a new far-red emitting fluorescent phosphoglycerolipid analogue
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DOI：
10.1039/c2fd20107k
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发表时间：
2013-01-01
影响因子：
3.4
通讯作者：
Eggeling, Christian
中科院分区：
文献类型：
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作者：
Honigmann, Alf;Mueller, Veronika;Eggeling, Christian
我们开发了一种明亮、光稳定、发射远红光的荧光磷酸甘油脂类似物，用于探测膜中脂质的扩散特性。该脂质类似物由饱和 (C18) 磷酸乙醇胺和亲水性远红发射荧光染料 (KK114) 组成，该染料通过长聚乙二醇接头连接到头部基团。与报道的远红发射荧光脂质类似物相反，该类似物主要划分为相分离三元双层的液体有序域。我们使用超分辨率 STED 显微镜 (STED-FCS) 进行荧光相关光谱分析，以测量新型脂质类似物在液体有序 (Lo) 和无序 (Ld) 相中的横向扩散。在云母载体上，我们观察微米大相，发现脂质类似物在所有测试的空间尺度（40-250 nm）上在Ld和Lo相中自由扩散，扩散系数分别为1.8 μ m(2) s(-1)和0.7 μ m(2) s(-1)。这表明Lo相的紧密分子堆积主要减缓扩散而不是引起异常的子扩散。沉积在酸洗玻璃上的相同三元混合物形成 Lo 纳米域
We have developed a bright, photostable, and far-red emitting fluorescent phosphoglycerolipid analogue to probe diffusion characteristics of lipids in membranes. The lipid analogue consists of a saturated (C18) phosphoethanolamine and a hydrophilic far-red emitting fluorescent dye (KK114) that is tethered to the head group by a long polyethylenglycol linker. In contrast to reported far-red emitting fluorescent lipid analogues, this one partitions predominantly into liquid ordered domains of phase-separated ternary bilayers. We performed fluorescence correlation spectroscopy with a super-resolution STED microscope (STED-FCS) to measure the lateral diffusion of the new lipid analogue in the liquid ordered (Lo) and disordered (Ld) phase. On a mica support, we observed micrometer large phases and found that the lipid analogue diffuses freely on all tested spatial scales (40-250 nm) in both the Ld and Lo phase with diffusion coefficients of 1.8 mu m(2) s(-1) and 0.7 mu m(2) s(-1), respectively. This indicates that the tight molecular packing of the Lo phase mainly slows down the diffusion rather than causing anomalous sub-diffusion. The same ternary mixture deposited on acid-cleaned glass forms Lo nano-domains of