Using Microcantilevers to Study the Interactions of Lipid Bilayers with Solid Surfaces

Abstract

We report the use of free-standing microcantilever beams, which have been used as an ultrasensitive method for measuring the surface free energy changes on a substrate induced by the adsorption of thin films, to probe the interactions between a solid surface and a phospholipid bilayer. We relate the observed deflection of a cantilever to the changes in the surface free energy of the solid surface which supports the phospholipid bilayer. We observe that the deflection is influenced by electrostatic and intermolecular interactions of the bilayer with the substrate. Increasing the surface charge density in the supported lipid bilayer (SLB), by increasing the ratio of cationic to zwitterionic lipids in bilayer, resulted in an increase in cantilever deflection. The surface free energy changes due to lipid transfer between anionic unilamellar vesicles and a cationic supported bilayer were also observed using microcantilevers. Finally, the adsorption free energy of a mixed lipid and cholesterol bilayer was measured demonstrating a detectable decrease in affinity between the phospholipid bilayer and the solid surface as a result of the addition of cholesterol. Our results reveal a new technique to probe the adsorption free energy of a SLB system as a function of the interactions governing the structure of supported lipid membranes.

Interaction of Extracellular Vesicles with Si Surface Studied by Nanomechanical Microcantilever Sensors

“…Substitution of this value into Equation (1) indicates that the surface stress exerted by the EVs adsorbed on the top Au face is higher than the one exerted by the EVs adsorbed onto the Si face by ∆σ = (28 ± 4) mJ/m 2 . This result can be discussed by taking into account the results reported in the literature about deflection signals upon injection of liposomes onto functionalized SiO 2 and Au surfaces [18]. As already discussed, Liu and co-workers demonstrated that upon incubation with POPC liposomes, a SLB forms on the SiO 2 surface, resulting in a positive deflection due to a compressive stress of ∆σ = (17.0 ± 1.4) mJ/m 2 .…”

“…By substituting ∆z into Equation (1) we learn that the layer formation involves a surface work that appears as a surface stress change of ∆σ = (8 ± 1) mJ/m 2 . This result is about half the value reported in the literature for the surface stress measured for the formation of supported lipid bilayer (SLB) on SiO2 surface from POPC liposomes [17,18]. This indicates that the formed layer is not continuous and/or that the formation of the EV layer is less energetic.…”

“…Experiments exploiting static deflection of MC beams are reported in literature to study the chemi-or physisorption of synthetic lipid bilayer on a cantilever surface, on either gold (Au) or silicon dioxide (SiO 2 ) surfaces [17,18]. Moreover, measurements of the resonance frequency shift induced upon phospholipid vesicle adsorption on oscillating cantilevers immersed in a liquid have been described [30].…”

“…Typical experiments exploit the well-known fact that liposomes readily adsorb and fuse on glass or silicon dioxide, giving rise to extended, homogeneous lipid double layers [45,46]. The common functionalization choice is then to protect the Au side of cantilevers and drive the vesicles fusion primarily on the SiO 2 surface [18]. Stability of liposomes on Au surfaces is instead not well understood: electrostatic (image-charge) attraction should provide strong adhesion leading to vesicles rupture and bilayer formation, yet adsorbed lipids exhibit negligible lateral diffusion.…”

“…The surface was selected as it is the most representative inorganic solid surface among the ones employed in biological applications, spanning from biosensors [14,15] to organoid-on-a-chip [16]. MC biosensors 2 of 11 were chosen for their ability to probe mass adsorption and interaction energetics at the solid-liquid interface of minute volumes of synthetic liposomes [17,18], comparable with standard EV formulations obtained from biological fluids or cell culture media, which are typically 100 µL solution volumes at few nM EV concentration (see for example [9]). Small EVs were isolated from an immortalized cell line of human vascular endothelial cell (HVEC) that is routinely used as model of EV-secreting cells, since it produces a good amount of highly pure exosomes compared to other cell types [19].…”