In order to study the influence on structural vibration from pre-stress and eccentricity, a experiment is designed. After analyzing the experimental results of 9 groups of modal tests on a post-tensioned unbonded pre-stress concrete beam, the following conclusions can be drawn: The influences on first order vibration frequency which are caused by eccentricity and pre-stress really exist. The rules of influences brought about by eccentricity and pre-stress on first order vibration frequency are complicated. When the eccentricity is close to the centroid, the linear correlation between the pre-stress and the first natural frequency will be good, on the contrary, the nonlinear will be strengthened. The first order vibrational frequency of the tested beam will go up with the increase of pre-stress.