Fluorescence methods for lipoplex characterization

AbstractSince the first reported transfection studies using cationic liposomes in 1987, significant advances have been made on the understanding of the physical properties of DNA/cationic liposome complexes (lipoplexes) in order to improve their transfection efficiencies. In this review a critical survey of the biophysical techniques used in their characterization is presented, with an emphasis on fluorescence methodologies, namely FRET. It is shown that the use of FRET combined with state-of-the-art modeling and data analysis allows detailed structural information in conditions close to the in vivo utilization of these non-viral based vectors. We describe in detail the use of fluorescence-based methods in (i) the assessment of DNA–lipid interaction and kinetics of lipoplex formation; (ii) membrane mixing studies; (iii) characterization of lipoplex molecular structure through the determination of interlamellar distances; and (iv) qualitative and quantitative evaluation of DNA condensation by cationic liposomes. This review aims at providing a framework for future characterization studies of novel liposomal formulations as gene delivery carriers, taking advantage of more sensitive nucleic acid and lipid dyes concomitantly with increasingly sophisticated fluorescence techniques