S100B levels may increase prior to a significant change in neurological function, or neuronal cell death. This is an important clinical finding inasmuch S100B levels in the normal range rule out cerebrovascular damage and BAY-60-7550 injury to the CNS in nearly 99% of patients by neurological imaging. Recently, controversy has arisen with regards to the brain specificity of this protein. Several lines of evidence suggest that extracerebral sources contribute to S100B serum levels. For example, in multi-organ system trauma without traumatic head injury, elevations in serum S100B levels collectively reflect traumatized fat, muscles, or bones since the blood-brain barrier remains intact in these situations and therefore, cannot account for the systemic rise in S100B levels. Similarly, shed blood from cardiac surgery displayed heightened levels without apparent head injury. Marathon runners, joggers, basketball players, ice hockey players, and boxers also have high S100B levels after engaging in their respective physical activity, which may be the result of both BBB opening and muscular release. The objective of this current study was to determine if tissue sources, outside the brain, contribute significantly to serum levels of S100B. In this study, we characterized by Western blot the expression of S100B in human tissue using two different antibodies and corroborated this data with mass spectrometry. The main contribution of this article to our understanding of S100B use in the clinical setting is that in spite of robust expression by extracranial sources, changes in serum levels are primarily dictated by extravasation across the disrupted BBB. In addition, we have shown that the main molecular species of total S100B related to blood-brain barrier disruption is the S100B homodimer. Such discrepancies may pose a significant problem, as they substantially impinge on and alter the conclusions drawn from experiments including those aiming at discoveries of disease treatment modalities or with diagnostic purpose. Another practical limitation of this study, due to the diversity and number of patients included, was the use of the BMI calculation to assess individuals’ relative body fat. Recent studies on nutrition and metabolism have validated techniques such as ultrasound, air displacement plethysmography and bioelectrical impedance to be superior to BMI for accurately measuring body fat. These techniques were not readily available nor could we easily implement them and therefore utilized the BMI calculation not only out of practicality, but also out of the widespread use of it in other S100B studies. Based on these observations, it may be desirable to incorporate any or all of the following to ascertain that an observed signal is indeed reflective of S100B: 1) Protein levels are generally caused by increased mRNA. Therefore, if release of S100B is due to a pathology initiating active synthesis rather than by passive release from necrotic astroglial cells quantitative RT-PCR of S100B messenger RNA would be appropriate to further support any observations.