We propose an on-chip cell manipulation method for trapping single motile cells in parallel. The proposed method traps large (≳50μm) motile cells in parallel, which is difficult to achieve by conventional cell manipulation methods based on optical, acoustic, electric, or magnetic forces. The trapping method exploits the flow induced by applying a vibration to a microfluidic chip with microstructures on its surface. By applying a rectilinear vibration to a chip with pairs of micropillars, we can trap single motile cells within the local flow generated between the micropillars. Using the proposed method, we trapped single Euglena gracilis cells (of size 50–100 μm) in parallel. Moreover, we evaluate the trapping performance for various micropillar array design parameters and the controllability of the trapping-flow velocity by varying the amplitude of the vibration. The proposed method was then demonstrated in a motility evaluation of motile cells. The demonstration confirms the potential of the proposed method in realizing high-throughput motility evaluations of single motile cells.