Hydrodynamic 3D flow-focusing techniques in microfluidics are categorized as (a) sheathless techniques which require high flow rates and long channels, resulting in high operating cost and high flow rates which are inappropriate for applications with flow rate limitations, and (b) sheath-flow based techniques which usually require excessive sheath flow rate to achieve hydrodynamic 3D flow-focusing. Many devices based on these principles use complicated fabrication methods to create multi-layer microchannels. We have developed a sheath-flow based microfluidic device that is capable of hydrodynamic 3D self-flow-focusing. In this device the main flow (black ink) in a low speed, and a sheath flow, enter through two inlets and enter a 180 degree curved channel (300 × 300 μm cross-section). Main flow migrates outwards into the sheath-flow due to centrifugal effects and consequently, vertical focusing is achieved at the end of the curved channel. Then, two other sheath flows horizontally confine the main flow to achieve horizontal focusing. Thus, the core flow is three-dimensionally focused at the center of the channel at the downstream. Using centrifugal force for 3D flow-focusing in a single-layer fabricated microchannel has been previously investigated by few groups. However, their demonstrated designs required high flow speed (>1 m/s) which is not suitable for many applications that live biomedical specie are involved. Here, we introduce a new design which is operational in low flow speed (<0.05 m/s) and is suitable for applications involving live cells. This microfluidic device can be used in detecting, counting and isolating cells in many biomedical applications.