Currently, there is no definitive diagnostic test for traumatic brain injury (TBI) to help physicians determine the seriousness of injury or the extent of cellular pathology. Calpain cleaves alphaII-spectrin into breakdown products (SBDP) after TBI and ischemia. Mean levels of both ipsilateral cortex (IC) and cerebral spinal fluid (CSF) SBDP at 2, 6, and 24 h after two levels of controlled cortical impact (1.0 mm and 1.6 mm of cortical deformation) in rats were significantly elevated by injury. CSF and IC SBDP levels were significantly higher after severe (1.6 mm) injury than mild (1.0 mm) injury over time. The correlation between CSF SBDP levels and lesion size from T2-weighted magnetic resonance images 24 hours after TBI as well as correlation of tau and S100beta was assessed. Mean levels of CSF SBDP (r = 0.833) and tau (r = 0.693) significantly correlated with lesion size while levels of CSF S100beta did not (r = 0.188). Although levels of CSF and IC SBDP and lesion size are all significantly higher after 1.6 mm than 1.0 mm injury, the correlation between CSF SBDP and lesion size was not significant following the removal of controls from the analysis. This indicates CSF SBDP is a reliable marker of the presence or absence of injury. Furthermore, larger lesion sizes 24 h after TBI were negatively correlated with motor performance on days 1-5 after TBI (r = -0.708). Based on these data, evaluation of CSF SBDP levels as a biomarker of TBI is warranted in clinical studies.