Nanoscale Characteristics of Ocular Lipid Thin Films Using Kelvin Probe Force Microscopy

Purpose: To describe the use of Kelvin probe force microscopy (KPFM) to investigate the electrical surface potential of human meibum and to demonstrate successful use of this instrument on both human meibum and a meibum model system (six-lipid stock [6LS]) to elucidate nanoscale surface chemistry and self-assembly characteristics. Materials and Methods: 6LS and meibum were analyzed in this study. Mica-supported thin films were created using the Langmuir-Blodgett trough. Topography and electrical surface potential were quantified using simultaneous atomic force microscopy/KPFM imaging. Results: Both lipid mixtures formed thin film patches on the surface of the mica substrate, with large aggregates resting atop. The 6LS had aggregate heights ranging from 41 to 153 nm. The range in surface potential was 33.0 to 125.9 mV. The meibum thin films at P = 5 mN/m had aggregates of 170 to 459 nm in height and surface potential ranging from 15.9 to 76.1 mV, while thin films at P = 10 mN/m showed an aggregate size range of 147 to 407 nm and a surface potential range of 11.5 to 255.1 mV. Conclusions: This study showed imaging of the differences in electrical surface potential of meibum via KPFM and showed similarities in nanoscale topography. 6LS was also successfully analyzed, showing the capabilities of this method for use in both in vitro and ex vivo ocular research. Translational Relevance: This study describes the use of KPFM for the study of ocular surface lipids for the first time and outlines possibilities for future studies to be carried out using this concept.