Nanoscale Wetting and Mechanical Characterization of Peptide Coacervates on Supported Lipid Bilayers by Atomic Force Microscopy

Peptide coacervates are dynamic, viscoelastic micro‐ and nanodroplets that selectively recruit diverse cargos and have emerged as a highly versatile platform for intracellular delivery, including for biomacromolecular therapeutics. Recent studies have focused on their cell uptake and intracellular cargo release, with growing interest in membrane interactions that regulate cell entry. However, most studies to date have relied on bulk techniques such as fluorescence recovery after photobleaching, microrheology, and quartz crystal microbalance, which offer ensemble‐averaged or indirect insights into droplet behavior. These approaches lack the resolution needed to directly assess how droplet wetting and mechanics influence interactions at biologically relevant interfaces. To overcome this limitation, a microfluidic atomic force microscopy platform is developed for single‐droplet measurements of wetting and elasticity on supported lipid bilayers. The method captures local mechanical and interfacial features by combining nm‐resolution topography with pixel‐resolved force mapping under biologically relevant conditions. HB peps , a peptide family with sequence‐defined mechanical tunability, form coacervates with broad variability in wetting and elastic modulus, modulated by peptide sequence, encapsulated cargo, and bilayer composition. These findings provide mechanistic insight into physical determinants governing coacervate–membrane interactions, identifying key parameters for rational design and establishing a new method for probing coacervate interactions at biologically relevant interfaces.