Direct Measurement of Charge Reversal on Lipid Bilayers using Heterodyne-Detected Second Harmonic Generation Spectroscopy

We report the detection of charge reversal induced by the adsorption of a cationic polyelectrolyte, poly(allylamine) hydrochloride (PAH), to buried supported lipid bilayers (SLBs), used as idealized model biological membranes. Through the use of an α-quartz reference crystal, we quantify the total interfacial potential at the interface in absolute units, using HD-SHG as an optical voltmeter in which the traditional wire leads of a voltmeter have been replaced by photons. This quantification is made possible by isolating from other contributions to the total SHG response the phase-shifted potential-dependent third-order susceptibility. We detect the sign and magnitude of the surface potential and the point of charge reversal at buried interfaces without prior information or complementary data. Isolation of the second-order susceptibility contribution from the overall SHG response allows us to directly characterize the Stern and Diffuse Layers over single-component SLBs formed from three different zwitterionic lipids of different gel-to-fluid phase transition temperatures (Tms). We determine whether the surface potential changes with the physical phase state (gel, transitioning, or fluid) of the SLB and incorporate 20 percent of negatively charged lipids to the zwitterionic SLB to investigate how the surface potential and the

second-order nonlinear susceptibility chi(2) change with surface charge.