Despite the tremendous advances in the understanding of the molecular mechanisms and the complexity of the diseases is one of the present challenges for the scientific community; then, novel strategies are required to be designed and developed for effective strategies for early diagnosis and treatment. As many cellular alterations are observed at protein level, high-throughput assays are dramatically needed for biomarker discovery. Herein, we describe advantages and limitations of protein microarrays, as proteomics strategy useful for multiplex and high-throughput protein characterization in clinical samples. Finally, a few examples are discussed; mostly of them related to currently disease biomarkers already identified in proximal fluids by protein arrays are discussed.