An actin filament is a micrometer-long biological filament, which serves as a track of transport systems in cells. The filament exhibits polarity while myosin transports a cargo along it unidirectionally. Here, we established a method to align many actin filaments parallel with each other on a substrate with uniform (>99%) polarity as assessed by myosin movements. This actin array is an ideal candidate for the construction of a nano-scale unidirectional transport system powered by myosin.