Redesigning solvatochromic probe Laurdan for imaging lipid order selectively in cell plasma membranes

Dmytro I Danylchuk , Erdinc Sezgin , Philippe Chabert , Andrey S Klymchenko (1)

1 LBP - Laboratoire de Bioimagerie et Pathologies

Imaging of biological membranes by environment-sensitive solvatochromic probes, such as Laurdan, provides information about organization of lipids, their ordering and uneven distribution. To address a key drawback of Laurdan linked to its rapid internalization and subsequent labelling of internal membranes, we redesigned it by introducing a membrane anchor group based on negatively charged sulfonate and dodecyl chain. The obtained probe, Pro12A, stains exclusively the outer leaflet of lipid bilayers of liposomes, as evidenced by leaflet-specific fluorescence quenching with a viologen derivative, and shows higher fluorescence brightness than Laurdan. Pro12A also exhibits stronger spectral change between liquid ordered and liquid disordered phases in model membranes and distinguishes better lipid domains in giant plasma membrane vesicles (GPMVs) than Laurdan. In live cells it stains exclusively the cell plasma membranes, in contrast to Laurdan and its carboxylate analogue C-Laurdan. Owing to its outer leaflet binding, Pro12A is much more sensitive to cholesterol extraction than Laurdan that redistributes within both plasma membrane leaflets and intracellular membranes. Finally, its operating range in the blue spectral region ensures absence of cross-talk with a number of orange/red fluorescent proteins and dyes. Thus, Pro12A will enable accurate multicolor imaging of lipid organization of cell plasma membranes in the presence of fluorescently-tagged proteins of interest, which will open new opportunities in biomembrane research.

Domaines: Chimie analytique

Dmytro I Danylchuk, Erdinc Sezgin, Philippe Chabert, Andrey S Klymchenko. Redesigning solvatochromic probe Laurdan for imaging lipid order selectively in cell plasma membranes. Analytical Chemistry, 2020, 92 (21), pp.14798-14805. ⟨10.1021/acs.analchem.0c03559⟩. ⟨hal-03086672⟩