title: The application of confocal microscopy to the study of liposome adsorption onto bacterial biofilms.
abstract: Confocal laser scanning microscopy has been used to visualise the adsorption of fluorescently labelled liposomes on immobilised biofilms of the bacterium Staphylococcus aureus. The liposomes were prepared with a wide range of compositions with phosphatidylcholines as the predominant lipids using the extrusion technique. They had weight average diameters of 125 +/- 5 nm and were prepared with encapsulated carboxyfluorescein. Cationic liposomes were prepared by incorporating dimethyldioctadecylammonium bromide (DDAB) or 3, beta [N-(N1,N1 dimethylammonium ethane)-carbamoyl] cholesterol (DC-chol) and anionic liposomes were prepared by incorporation of phosphatidylinositol (PI). Pegylated cationic liposomes were prepared by incorporation of DDAB and 1,2-dipalmitoylphosphatidylethanolamine-N-[polyethylene glycol)-2000]. Confocal laser scanned images showed the preferential adsorption of the fluorescent cationic liposomes at the biofilm-bulk phase interface which on quantitation gave fluorescent peaks at the interface when scanned perpendicular (z-direction) to the biofilm surface (x-y plane). The biofilm fluorescence enhancement (BFE) at the interface was examined as a function of liposomal lipid concentration and liposome composition. Studies of the extent of pegylation of the cationic liposomes incorporating DDAB, on adsorption at the biofilm-bulk phase interface were made. The results demonstrated that pegylation inhibited adsorption to the bacterial biofilms as seen by the decline in the peak of fluorescence as the mole% DPPE-PEG-2000 was increased in a range from 0 to 9 mole%. The results indicate that confocal laser scanning microscopy is a useful technique for the study of liposome adsorption to bacterial biofilms and complements the method based on the use of radiolabelled liposomes.
authors.affiliation: School of Biological Sciences, University of Manchester, Manchester, M13 9PL, UK.
authors.forename: Khalid
authors.initials: K
authors.lastname: Ahmed
authors.forename: Phillip N
authors.initials: PN
authors.lastname: Gribbon
authors.forename: Malcolm N
authors.initials: MN
authors.lastname: Jones
mesh_terms: D000327:Adsorption; D018441:Biofilms; D005452:Fluoresceins; D005453:Fluorescence; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism*; D018613:Microscopy, Confocal / Q000379:methods*; D010713:Phosphatidylcholines; D013211:Staphylococcus aureus / Q000254:growth & development* / Q000378:metabolism*
chemical_list: D005452:Fluoresceins; D008081:Liposomes; D010713:Phosphatidylcholines; C024098:6-carboxyfluorescein