Membrane fluidity is a critical parameter of cellular membranes, which cells continuously strive to maintain within a viable range. Interference with the correct membrane fluidity state can strongly inhibit cell function. Triggered changes in membrane fluidity and associated impacts on lipid domains have been postulated to contribute to the mechanism of action of membrane targeting antimicrobials, but the corresponding analyses have been hampered by the absence of readily available analytical tools. Here, we expand upon the protocols outlined in the first edition of this book, providing further and alternative protocols that can be used to measure changes in membrane fluidity. We provide detailed protocols, which allow straightforward in vivo and in vitro measurement of antibiotic compound-triggered changes in membrane fluidity and fluid membrane microdomains. Furthermore, we summarize useful strains constructed by us and others to characterize and confirm lipid specificity of membrane antimicrobials directly in vivo.
Keywords: DiIC12; Fatty acid disorder; Laurdan; Lipid domains; Lipid mutants; Lipid packing; Membrane fluidity; Membrane microdomains; Membrane targeting antimicrobials; Nile Red; RIFs.
© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.