Zirconium ion mediated formation of liposome multilayers

Phospholipid vesicles have attracted considerable interest as a platform for a variety of biomolecular binding assays, especially in the area of membrane protein sensing. The development of liposome-based biosensors widely relies on the availability of simple and efficient protocols for their surface immobilization. We present a novel approach toward the creation of three-dimensional phospholipid vesicle constructs using multivalent zirconium ions as linkers between the liposomes. Such three-dimensional sensing platforms are likely to play a key role in the development of biosensing devices with increased loading capacity and sensitivity. After demonstrating the affinity of Zr(4+) toward the phospholipids, we formed vesicle multilayers by sequential injections of solutions containing either liposomes or ZrOCl(2). In situ adlayer characterization was carried out by optical waveguide lightmode spectroscopy (OWLS) and quartz crystal microbalance with dissipation (QCM-D) measurements while imaging was performed by atomic force microscopy (AFM) and fluorescence microscopy. Multilayers were successfully constructed, and as demonstrated in a model fluorescence-based biomolecular binding assay, the sensor's loading capacity was increased. Furthermore, we observed that lipid exchange between the vesicles is promoted in the presence of Zr(4+) and that addition of a phosphate-containing buffer leads to adlayer loosening and creation of lipidic tubular structures. The approach presented here could be applied to the study of membrane proteins in a highly sensitive manner due to the increased surface area or to produce functional coatings for controlled drug release and host response.