In vivo, tissue-level, mechanical thresholds for axonal injury in the guinea pig optic nerve were determined by comparing morphological injury to estimated in vivo tissue strain. The right optic nerve of adult male guinea pigs was stretched to one of seven ocular displacement levels. Morphological injury was detected three days post-stretch with neurofilament immunohistochemical staining (NF68). A companion set of in situ experiments was used to determine the empirical relationship between ocular displacement and optic nerve stretch. Logistics regression analysis, combined with sensitivity and specificity measures and receiver operating characteristic (ROC) curves were then used to predict strain thresholds for axonal injury.From this analysis, we determined three Lagrangian strain- based thresholds for morphological damage to the guinea pig white matter. The threshold intended to minimize the false positive rate was a strain of 0.34, and the threshold that minimized the false negative rate was 0.14. The threshold criteria that balanced the specificity and sensitivity measures was 0.21. With this threshold data, one may gain new insights into the occurrence of DAI in man.