Abstract
Microdomains corresponding to localized partition of lipids between ordered and less ordered environments are the subject of intensive investigations, because of their putative participation in modulating cellular responses. One popular approach in the field consists in labelling membranes with solvatochromic fluorescent probes such as laurdan and C-laurdan. In this report, we describe a high-yield procedure for the synthesis of laurdan, C-laurdan and two new fluorophores, called MoC-laurdan and M-laurdan, as well as their extensive photophysical characterization. We find that the latter probe, M-laurdan, is particularly suited to discriminate lipid phases independently of the chemical nature of the lipids, as measured by both fluorescence Generalized Polarization (GP) and anisotropy in large unilamellar vesicles made of various lipid compositions. In addition, staining of live cells with M-laurdan shows a good stability over time without any apparent toxicity, as well as a wider distribution in the various cell compartments than the other probes.
Highlights
Numerous physiological processes take place at the cell plasma membrane and its organization into domains participates in modulating many cellular responses[1]
M-laurdan, MoC-laurdan and C-laurdan on LUVs made of DPPC, DPPC/cholesterol (6:4) and POPC, we performed Generalized Polarization (GP) and anisotropy measurements to compare both types of samples and we found no detectable difference, suggesting that the presence of low concentrations of dimethyl sulfoxide (DMSO) in the samples does not have any significant effect on this kind of probes (Figure S4)
When incorporated into a DPPC bilayer, all four probes showed a steep decrease for both GP and anisotropy when heated above the melting transition of 41°C, indicating that the four probes are equivalently sensing the phase behavior from solid like phase (So) to liquid disordered phase (Ld) occurring in the bilayer, in agreement with the results found in the literature for laurdan[43]
Summary
Numerous physiological processes take place at the cell plasma membrane and its organization into domains participates in modulating many cellular responses[1]. The external leaflet of plasma membranes (PMs) is mainly composed of phosphatidylcholine, sphingomyelin and cholesterol while the inner leaflet contains significant amounts of phosphatidylserine and phosphatidylethanolamine[2]. Pure sphingomyelin is known to form solid like phase (So) at physiological temperature[5], due to its long acyl chain. In cell membranes containing cholesterol, cholesterol and sphingomyelin have been shown to interact and form liquid ordered phases (Lo) surrounded by a liquid disordered phase (Ld) essentially comprised of phosphatidylcholine[2]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
