COMPARATIVE ANALYSIS OF MEMBRANE LIPID ORDER IMAGING WITH TWO FLUORESCENT PROBES IN INTACT CELLS
RALITSA VELEVA1,2*, GALYA STANEVA2, TANYA TOPOUZOVA-HRISTOVA1
1 – Faculty of Biology, Sofia University “St. Kliment Ohridski”, Sofia, Bulgaria
2 – Institute of Biophysics and Biomedical Engineering, BAS, Sofia, Bulgaria
*Corresponding author: a._k.c@abv.bg
Keywords: cell membranes, lipid rafts, fluorescence probes, fluorescence microscopy
Abstract: Cell membranes include vast range of lipids which form segregated regions of structure and function. They assemble small, dynamic clusters of lipids in a more ordered state called “rafts”. These microdomains mediate cellular processes such as protein sorting, signal transduction, membrane trafficking and cell adhesion. Large part of our knowledge about the behaviour of lipid mixtures in bilayers is shaped from studies on model membranes which are with well-defined composition. Our aim was to observe domains on the more complex membrane systems of intact cells.
We applied membrane probes which give information on the degree of lipid packing to make membrane domains visible. We compared two polarity sensitive fluorescent probes – namely Laurdan and di-4-ANEPPDHQ. Two cell lines with distinct differences in their plasma membrane composition and two fluorescent methods for analysis of rafts – microscopy and spectroscopy were chosen. We obtained reliable assessment of the state of the membrane order through both methods. Di-4-ANEPPDHQ provided better micrographs with the available microscope technique.