S-100 protein, described initially by Moore, constitutes a large family of at least 20 proteins with calcium binding ability. It is found as homo- or hetero-dimers of two different subunits (A and B). Types S-100AB and S-100BB are described as S-100B protein and are shown to be highly specific for nervous tissue. It is present in the cytosol of glial and Schwann cells, and also in adipocytes and chondrocytes, although in very low concentrations in the latter two. The role of protein S-100B is not yet fully understood. It is suggested that it has intracellular and extracellular neurotropic as well as neurotoxic function. At nanomolar levels, S-100B stimulates neurite outgrowth and enhances survival of neurons. However, at micromolar levels it stimulates the expression of inflammatory cytokines and induces apoptosis. Recently, serum S-100B protein has been proved to be an attractive surrogate marker of primary severe brain injury and secondary insults. It can be measured in the arterial and venous serum; it is not affected by haemolysis and remains stable for several hours without the need for immediate analysis. Its short half-life makes measurements crucial in the emergency and intensive care settings. This review summarises published findings on S-100B regarding its role as a serum biochemical marker of brain injury, i.e., after severe, moderate or mild neuro-trauma, subarachnoid haemorrhage, thrombo-embolic stroke, cerebral ischaemia and brain tumours, as well as extracranial trauma, neurodegenerative and psychiatric disorders.