Since most conventional clinical assays are performed on cell-free serum or plasma, plasma isolation is a necessary step in the processing of whole blood sample pretreatment. Traditional technologies for plasma isolation are based on centrifugation or membrane filtration, which are time and cost intensive processes. Microfluidic chips for whole blood plasma isolation are attractive alternatives, which lead to more gentle, fast, and consistent manipulation of the whole blood samples, and therefore more accurate and better quality of plasma isolation. In this chapter, a variety of microfluidic devices for blood plasma separation are reviewed and compared. Two specific examples of plasma isolation leveraging MEMS technologies are included for further concept demonstration in which design, fabrication and testing of these microfluidic devices are covered in detail. These plasma isolation microfluidic devices can be easily connected with other microfluidic components to form micro total analysis systems for blood testing with the advantages of continuous and real time separation of plasma from whole blood samples.