This Account describes a new paradigm, Langmuir-Blodgett (LB) patterning, for large-area patterning with mesostructured features based on the well-established LB technique. This strategy uses a simple fabrication technique to control the alignment, size, shape, and periodicity of self-organized phospholipid monolayer patterns with feature sizes down to 100 nm over surface areas of square centimeters. Because of the anisotropic wetting behavior of the patterns, they can be used as templates to direct the self-assembly of functional molecules and nanocrystals. Furthermore, the chemical patterns can be converted into topographical structures, which can be used to direct cell growth and organize nanocrystals. The mesoscopic structured surfaces described here may serve as a platform in engineering the biological/material interface and constructing biofunctionalized structures and "programmed" systems.