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Iron is an essential element to the well functionning of the organism and requires careful maintenance of its homeostasis. This is mainly due to hepcidin, a hormone secreted by the liver that controls the flow of iron within the body. It has a hyposideremic action by reducing the expression of ferroportin, the only protein known to this day, which can export iron into the extracellular environment. This has the effect of decreasing intestinal absorption and increasing intracellular retention, especially in macrophages. Hepcidin is stimulated by elevation of iron and inflammation while activation of erythropoiesis inhibits it. Understanding its regulation allows a better understanding of the pathophysiological mechanisms of overload diseases and iron deficiency. Therefore, hepcidin analysis is interesting for the exploration of iron homeostasis. In combination with other biological parameters of iron status, it is possible to find better instructions for therapeutic managements of iron metabolism diseases. Thus, an assaying method by coupling liquid chromatography and tandem mass spectrometry has been implemented at Grenoble University Hospital and the analytical performances of this assay met the laboratory requirements in terms of reliability of the analysis.