This paper proposes a high speed cell stiffness evaluation with high reliability. The procedure includes both online measurement and offline inspection. By using the property where the passing time of the cell in the micro channel is a function of the cell stiffness and its size, a real time vision system whose handling frequency is 6 kHz can capture the size as well as the passing time of each cell. During the online measurement, we save two image data at both the entrance and the exit for each cell. In offline manual inspection, all doubtful data are killed so that we can make sure the reliability. Experimental results ensure that the proposed approach can handle 500 cells within 10min in including both online and offline procedures.