Cell separation is used to separate homogeneous and individual cell classes from a heterogeneous cell population. The efficiency and purity of these separated cells are of great importance in personalized medicine, regenerative medicine, disease monitoring and drug testing as well as in the therapeutic and diagnostic research. In this thesis, different microfluidic approaches were presented for cell separation. With this regard, a closed channel vacuum-integrated microfluidic chip was developed using an air permeability of a Polydimethylsiloxane and density-based separation of microparticles was performed. Besides, a centrifugal microfluidic system, Spinochip, was developed with one reservoir as inlet and outlet for the first time and different fluid manipulations were shown in the system. The system was applied to clinical tests of hematocrit measurements and white blood cell estimation using real patient samples. The developed system offered correlated results with clinical results. In addition to closed channel microfluidics, negative-magnetophoresis microfluidic chip was demonstrated for the size-based separation of microparticles and cells. In this regard, capturing rate of breast cancer cells (MCF-7) and human monocyte cells (U937) was investigated. The results showed that the approaches presented here could promote to the microfluidic studies for size-based cell separation.