The manipulation of particles and cells in micro-fluids, such as cell suspensions, is a fundamental task in Lab-on-a-Chip applications. According to their analysis purposes in either the pre- or post-processing stage, particles/cells flowing inside a microfluidic channel are handled by means of enriching, trapping, separating or sorting. In this study, we report the use of patterning flows produced by a series of grooved surfaces with different geometrical setups integrated into a microfluidic device, to continuously manipulate the flowing particles (5 to 20 μm in diameters) of comparable sizes to the depth of the channel in ways of: 1) concentrating, 2) focusing, and 3) potential separating. The device is fabricated using soft lithographic techniques and is composed of inlets, microfluidic channels, and outlets for loading, manipulating and retrieving cell suspensions, respectively. Such fabrication methods allow rapid prototyping of micron or submicron structures with multiple layers and replica molding on those fabricated features in a clear polymer. The particles are evenly distributed in the entrance of the microchannel and illustrate the enriching, focusing, or size-selective profiles after passing through the patterning grooves. We expect that the techniques of manipulating cell suspensions from this study can facilitate the development of cell-based devices on 1) the visualization of counting, 2) the visualization of sizing, and 3) the particle separating.