Influence of borate eluent pH value and competition eluant on regeneration of the used IMAC-Cu proteinchip is investigated by SEM and SELDI-TOF-MS proteinchip detection system. The results show that proteins are easier to be removed from the used proteinchip surface with the decreasing of borate eluent pH value, but its destruction to the chemical structure of the proteinchip also grows in intensity. The elution effect of protein on IMAC-Cu proteinchip can be significantly enhanced when the competition eluant Gly or EDTA is added to borate eluate. However, comparing with Gly, EDTA as competition eluant has generation effect. The absorption protein quantity and type of the regenerated IMAC-Cu proteinchip by the pH 8 borate eluent added 0.02mol/L EDTA for 72h is very similar to the fresh chip after absorbing same serum of cancer patients.