An antireflective optical film with subwavelength structures replicated by use of a roll-to-roll micro-replication process (RMRP) is investigated. Firstly, a single layer of a nanostructure on a polymer film is designed for an antireflection purpose by the finite difference time domain method in the visible light spectrum. Structures of a conical cylinder array, with spatial period of 400 nm, diameter of 200 nm and height of 350 nm, are numerically obtained. Then, such structures are fabricated by RMRP combining originated structure fabrication realized by deep ultra-violet lithography and dry etching, Ni mold electroplating and replication by using the roll-to-roll process imprinting into the flexible polyethylene terephthalate substrate. A nanostructure roller mold bonded with Ni molds has been successfully fabricated and coated with the self-assembly monolayer process for the purpose of fabricating an anti-adhesion film and improving the lifetime of the Ni molds. The duplicated nanostructure films show a good optical quality of antireflection (AR ≤ 2.45% in a 400-700 nm spectral range) and are in good agreement with the theoretical predictions. The experimental results show that the developed process is a promising and cost-effective method for the continuous duplication of flexible devices with nano-scaled feature sizes used in nanophotonics by RMRP.