The micro-molding of thermoplastic polymer is a popular fabrication method for micro/nano devices because several thousand parts can be produced after a mold insert has been made. Most micro-molding techniques such as hot embossing or injection molding use thermal energy to plasticize the thermoplastic polymer, and require an additional cooling stage. Recently, ultrasonic vibration energy has been used to soften a local region of thermoplastic polymer by inducing frictional heat between an ultrasonic tool horn and the polymer material. The present study aims to develop an ultrasonic patterning process in which the micro-patterns engraved on the tool horn are replicated onto the surface of a polymer film. For this purpose, the design of a tool horn for the ultrasonic patterning was investigated to obtain good replication characteristics of micro-patterns with uniformity thorough the entire pattern area. Finite element analyses were conducted to predict the vibration characteristics of the tool horn. Statistical analysis using the response surface method was concurrently performed to find the optimal design conditions to ensure high and uniform pattern replicability. Experimental verifications were then followed by application of the proposed horn design.