This report presents the fabrication of a micro fluidic device using the UV lithography method in order to separate nonmagnetic fluoresbrite carboxy microspheres from the mixed ferrofluids in microchannels. The microfluidic device is fabricated using a SU8-50 negative photoresist that is exposed to the UV lights with a mask (Y shape) on the top of the photoresist. By applying the external magnet from a side of the microchannel, the fluoresbrite carboxy microspheres and ferrofluids are separated into different channels because of the magnetic force acted on the nonmagnetic spheres. During the fabrication, a number of different parameters, such as UV exposure times, UV power, photoresist thickness, etc. were conducted and optimized for our needs. In addition, in the magnetic field testing, different pumping speeds, and particle concentrations associated with the different distances between the magnet and the microfluidic system were studied for an efficient separation.