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A multitude of biological processes are enabled by complex interactions between lipid membranes and proteins. To understand such dynamic processes, it is crucial to differentiate the constituent biomolecular species and track their individual time evolution without invasive labels. Here, we present a label-free mid-infrared biosensor capable of distinguishing multiple analytes in heterogeneous biological samples with high sensitivity. Our technology leverages a multi-resonant metasurface to simultaneously enhance the different vibrational fingerprints of multiple biomolecules. By providing up to 1000-fold near-field intensity enhancement over both amide and methylene bands, our sensor resolves the interactions of lipid membranes with different polypeptides in real time. Significantly, we demonstrate that our label-free chemically specific sensor can analyze peptide-induced neurotransmitter cargo release from synaptic vesicle mimics. Our sensor opens up exciting possibilities for gaining new insights into biological processes such as signaling or transport in basic research as well as provides a valuable toolkit for bioanalytical and pharmaceutical applications.

In this work, we present an infrared plasmonic biosensor for chemical-specific detection and monitoring of biomimetic lipid membranes in a label-free and real-time fashion. Lipid membranes constitute the primary biological interface mediating cell signaling and interaction with drugs and pathogens. By exploiting the plasmonic field enhancement in the vicinity of engineered and surface-modified nanoantennas, the proposed biosensor is able to capture the vibrational fingerprints of lipid molecules and monitor in real time the formation kinetics of planar biomimetic membranes in aqueous environments. Furthermore, we show that this plasmonic biosensor features high-field enhancement extending over tens of nanometers away from the surface, matching the size of typical bioassays while preserving high sensitivity.

During vesicle budding or endocytosis, biomembranes undergo a series of lipid- and protein-mediated deformations involving cholesterol-enriched lipid rafts. If lipid rafts of high bending rigidities become confined to the incipient curved membrane topology such as a bud-neck interface, they can be expected to reform as ring-shaped rafts. Here, we report on the observation of a disk-to-ring shape morpho-chemical transition of a model membrane in the absence of geometric constraints. The raft shape transition is triggered by lateral compositional heterogeneity and is accompanied by membrane deformation in the vertical direction, which is detected by height-sensitive fluorescence interference contrast microscopy. Our results suggest that a flat membrane can become curved simply by dynamic changes in local chemical composition and shape transformation of cholesterol-rich domains.

During vesicular trafficking and release of enveloped viruses, the budding and fission processes dynamically remodel the donor cell membrane in a protein- or a lipid-mediated manner. In all cases, in addition to the generation or relief of the curvature stress, the buds recruit specific lipids and proteins from the donor membrane through restricted diffusion for the development of a ring-type raft domain of closed topology. Here, by reconstituting the bud topography in a model membrane, we demonstrate the preferential localization of cholesterol- and sphingomyelin-enriched microdomains in the collar band of the bud-neck interfaced with the donor membrane. The geometrical approach to the recapitulation of the dynamic membrane reorganization, resulting from the local radii of curvatures from nanometre-to-micrometre scales, offers important clues for understanding the active roles of the bud topography in the sorting and migration machinery of key signalling proteins involved in membrane budding.

We integrate periodic nanohole arrays in a thin gold film with lipid membranes in a microfluidic channel. Surface plasmon-enhanced light transmission through the periodic nanohole arrays enables real-time label-free sensing of molecular binding on the lipid membrane surface. This membrane biosensor can potentially act as a natural platform for studying binding kinetics of proteins with their binding partners anchored in the lipid membrane. We also present the concept of using nanopore arrays for kinetic assays of transmembrane proteins in a free-standing lipid membrane.

Highlights • Electrodes with 20 nm-wide gaps were fabricated by atomic layer lithography. • Nanogap electrodes enable low-voltage dielectrophoresis (DEP) trapping. • Spherical supported lipid bilayers and myelin particles were trapped with DEP. • Integrated microfluidics allow buffer switching and reagent delivery for binding assays. We demonstrate a gold nanogap electrode platform that can rapidly create a linear array of biological particles by low-voltage dielectrophoresis (DEP). We further combine microfluidic buffer exchange to introduce protein molecules in high-conductivity solutions while trapping and immobilizing particles. The nanogap between the gold electrodes enables low operating voltages that prevent unwanted Joule heating in high-conductivity buffers. This platform is used to trap bioparticles composed of lipid membranes such as spherical supported lipid bilayers and brain-derived myelin particles, followed by detection of protein binding to specific membrane-bound receptors. We use bioparticles with different sizes, physicochemical properties, and origins to demonstrate a platform that can be used to study a variety of biomolecular interactions. The low-power linear DEP trap combined with microfluidic buffer exchange has potential to enable a portable biosensing platform to rapidly concentrate rare biological particles and perform on-chip binding assays with improved detection limits in physiological buffers. [ABSTRACT FROM AUTHOR]

A label-free, non-dispruptive, and real-time analytical device to monitor the dynamic features of biomolecules and their interactions with neighboring molecules is an essential prerequisite for biochip- and diagonostic assays. To explore one of the central questions on the lipid-lipid interactions in the course of the liquid-ordered (l o) domain formation, called rafts, we developed a method of reconstituting continuous but spatially heterogeneous lipid membrane platforms with molayer-bilayer juntions (MBJs) that enable to form the l o domains in a spatiotemporally controlled manner. This allows us to detect the time-lapse dynamics of the lipid-lipid interactions during raft formation and resultant membrane phase changes together with the raft-associated receptor-ligand binding through the surface plasmon resonance (SPR). For cross-validation, using epifluorescence microscopy, we demonstrated the underlying mechanisms for raft formations that the infiltration of cholesterols into the sphingolipid-enriched domains plays a crucial roles in the membrane phase-separation. Our membrane platform, being capable of monitoring dynamic interactions among lipids and performing the systematic optical analysis, will unveil physiological roles of cholesterols in a variety of biological events. Continuous but spatially heterogeneous membranes were probed by the SPR for monitoring the kinetic behavior of lipid-lipid interactions (a) and the accompanying biochemical activity of ligand-receptor binding. Our monolayer-bilayer junction methodology of manipulating the bio-membranes enables to precisely control the phase-separation of l o domains at pre-defined regions (b) on a SPR-based biochip and to unveil the kinetics of the raft formation together with raft-associated receptor-ligand interactions (c). Image 1 • Continuous but spatially heterogeneous membranes were probed by the SPR for monitoring the kinetic behavior of lipid-lipid interactions • The accompanying biochemical activity of ligand-receptor binding. Our monolayer-bilayer junction methodology of manipulating the bio-membranes enables to precisely control the phase-separation of l o domains at pre-defined regions on a SPR-based biochip and to unveil the kinetics of the raft formation together with raft-associated receptor-ligand interactions [ABSTRACT FROM AUTHOR]