This paper reports a simple and rapid method for fabrication of microfluidic chips on polymethylmethacrylate (PMMA) substrate using a flexible and low-cost CO2 laser system. The CO2 laser employed has a wavelength of 10.6 mu m. The laser power used for channel fabrication ranged from 3 and 12 W, the beam travel speeds ranged from 5 to 50 mm/s and the passes were varied in the range of 1 to 3 times. Typical channel depths were between 100 and 900 mu m, while the width of fabricated channels ranged from 100 to 300 mu m. The effects of the process parameters (the laser power, the beam travel speed of the laser beam and the number of passes) on the dimensional quality (the depth, the width and their aspect ratio) of the microchannel manufactured from PMMA were experimentally investigated. The change law of the channel geometry depending on process parameters was obtained. A high reproducibility of micro-channel geometry was attained. At last, a CO2 laser output power of 5.5 W and a laser beam travel speed of 35 mm/s combining a hot press bonding technique were chosen to fabricate a microfluidic chip within half hour. The pattern qualities and experimental results confirm that the CO2 laser micromachining technology has a great potential for application in flexible, rapid and economic production of polymeric microfluidic chips.