Protein profiling with high-throughput proteomic technology SELDI-TOF is a new potential tool for diagnostic of human analytical reproducibility is a significant challenge in SELDI-TOF profiling in order to have confidence in the results. Here we showed a simple step to improve its analytical performance. Incubation of denaturated samples overnaight at 4 deg C increased significantly reproducibility and quality of proteomic profile of SELDI-TOF MS for IMAC30-Cu ProteinChip. This strategy could be concerned and apply to address reproducibility issue in this system.