A method based on co-flow microfluidic device and photoinitiated polymerization was applied to prepare polymethyl methacrylate (PMMA) microspheres with controllable and uniform particle size. The effects of the quantity of polyvinyl alcohol (PVA), sodium dodecyl sulfate (SDS), ethylene glycol, flow rate of continuous phase, capillary tip diameter and necking degree of outer tube on the particle size and uniformity of microspheres were investigated. The morphology of microspheres was observed by microscope, and the diameters of microspheres were measured. The results show that the diameter of microspheres decreases with the increase of the quantity of PVA or SDS. And the particle size distribution of microspheres changes with the variation of PVA. The effect of ethylene glycol on the particle size of the microspheres is not significant. And the increase of the flow rate of continuous phase or necking degree of the outer tube, or the decrease of capillary tip diameter will lead the decrease of the particle size of the prepared microspheres. The size of the prepared microspheres ranges from 65μm to 316 μm, and the coefficient of variation (CV) can be controlled less than 3%. This paper provides a method for the controllable preparation of PMMA microspheres and other polymer microspheres with different sizes and uniform particle sizes.