Microfluidic devices having an
electrowetting configuration and an optimized droplet actuation surface are provided. The devices include a conductive substrate having a
dielectric layer, a hydrophobic layer covalently bonded to the
dielectric layer, and a first
electrode electrically coupled to the
dielectric layer and configured to be connected to a
voltage source. The microfluidic devices also include a second
electrode, optionally included in a cover, configured to be connected to the
voltage source. The hydrophobic layer features self-associating molecules covalently bonded to a surface of the
dielectric layer in a manner that produces a densely-packed
monolayer that resists intercalation and or penetration by polar molecules or species. Also provided are microfluidic devices having an
electrowetting configuration that further include a section or module having a
dielectrophoresis configuration; systems that include any of the microfluidic devices in combination with an
aqueous droplet and a fluidic medium immiscible with the medium of the
aqueous droplet; related kits; and methods of manipulating droplets, optionally containing micro-objects such as biological cells, within the microfluidic devices.