Microfluidic devices are highly commonplace in the field of biomedical technology, point of care diagnostics and chemical analysis. The rapid and low cost manufacturing of these devices have always been a challenge. CO2 laser micromachining has played an important role in micro-machining of devices at a scale similar to the microfluidic devices although it renders the machined surfaces with high surface roughness. The chapter reports an initiative to do process optimization of laser micromachining technique for producing smooth machined surfaces in the micro scale devices. The chapter discusses the impact of process parameters like raster speed, laser power, print resolution etc. and its optimization using two target functions of dimensional precision and surface roughness on micro-channels made in PMMA (Poly methyl metha acrylate) substrates. The laser machined PMMA samples are analyzed using 3D-profilometry and Field emission scanning electron microscope (FESEM) for surface quality and dimensional precision. To investigate optimum process parameters of CO2 laser for fabricating the micro-channel on PMMA with dimensional accuracy and good surface quality, Analysis of variance (ANOVA) and regression analysis is conducted. It is found that optimum surface roughness of this process is around 7.1 µm at the optimum value of the process parameters 7.5 mm/s (50 % of maximum machine limit) raster speed, 17.9 W (51 % of maximum machine limit) laser power and 1200 DPI (100 % of maximum machine limit) printing resolution. The static contact angle of the micro-machined surface has also been observed for analyzing the amenability of these channels to flow of water like fluids for micro-fluidic applications. The chapter also covers a review of work done by various researchers in which they developed different methodology for successful manufacturing of microfluidic devices by employing CO2 laser micromachining. © 2015, Springer India.