Abstract
The lateral organization of molecules in the cellular plasma membrane plays an important role in cellular signaling. A critical parameter for membrane molecular organization is how the membrane lipids are packed. Polarity-sensitive dyes are powerful tools to characterize such lipid membrane order, employing, for example, confocal and two-photon microscopy. The investigation of potential nanodomains, however, requires the use of superresolution microscopy. Here, we test the performance of the polarity-sensitive membrane dyes Di-4-ANEPPDHQ, Di-4-AN(F)EPPTEA, and NR12S in superresolution stimulated emission depletion microscopy. Measurements on cell-derived membrane vesicles, in the plasma membrane of live cells, and on single virus particles, show the high potential of these dyes for probing nanoscale membrane heterogeneity.
Highlights
The lateral organization of molecules in the cellular plasma membrane has significant influence on cellular functions
Such properties have been indicated for recently developed probes such as Di-4-ANEPPDHQ, Di-4-AN(F)EPPTEA, and NR12S [17,28,29,30,32,33] (Fig. 1)
We have shown the compatibility of the polarity-sensitive membrane dyes Di-4-ANEPPDHQ, Di-4-AN(F)EPPTEA, and NR12S for superresolution stimulated emission depletion (STED) microscopy
Summary
The lateral organization of molecules in the cellular plasma membrane has significant influence on cellular functions. Lipid-lipid and lipid-protein interactions facilitate the segregation of plasma membrane molecules into clusters or nanodomains that constitute catalytic platforms for a myriad of activities such as cellular signaling [1,2]. Lateral heterogeneity in lipid order, i.e., how the packing of lipids varies over space, and how this is involved in molecular segregation, is of interest because membrane order may modulate protein functionality [3,4,5,6,7]. Certain lipid combinations (especially those involving cholesterol) result in macroscopic phase separation in model membranes, which can readily be resolved by confocal microscopy [8]. Methodological advances are needed to adequately tackle this phenomenon of high biological relevance
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