S100A8 and S100A9 (also known as MRP8 and MRP14, respectively) are Ca binding proteins belonging to the S100 family. They often exist in the form of heterodimer, while homodimer exists very little because of the stability. S100A8/A9 is constitutively expressed in neutrophils and monocytes as a Ca sensor, participating in cytoskeleton rearrangement and arachidonic acid metabolism. During inflammation, S100A8/A9 is released actively and exerts a critical role in modulating the inflammatory response by stimulating leukocyte recruitment and inducing cytokine secretion. S100A8/A9 serves as a candidate biomarker for diagnosis and follow-up as well as a predictive indicator of therapeutic responses to inflammation-associated diseases. As blockade of S100A8/A9 activity using small-molecule inhibitors or antibodies improves pathological conditions in murine models, the heterodimer has potential as a therapeutic target. In this review, we provide a comprehensive and detailed overview of the distribution and biological functions of S100A8/A9 and highlight its application as a diagnostic and therapeutic target in inflammation-associated diseases.