Roads represent a serious obstacle to maintaining ecological connectivity and viable wildlife populations. Wildlife crossings have been designed and incorporated into road construction projects to mitigate these impacts. The 24 wildlife crossings in Banff National Park have been the subject of intensive ecological studies since 1996. Currently more than 50,000 passes by 10 species of large mammals have been recorded. The actual number of individuals using the passages is unknown and only rough estimates can be made. Further, up until now assessments of mitigation effectiveness have been focused at the level of individuals and not populations. Healthy functioning ecosystems require viable wildlife populations. Thus, it is critical to know the performance of these measures at the population level. Central to determining the utility of wildlife crossings in enhancing population viability is knowing what proportion of individuals of a population are using the passages. At present, the most reliable method is live-trapping and mark-recapture analysis, but for logistical reasons this is impractical. Recent developments in molecular techniques allow for species-specific DNA from hair samples, thus making it possible to identify individual animals, their sex, and genetic relatedness.This pilot study will test techniques and develop a protocol for systematically sampling and genotyping of hairs “captured” from passing animals at wildlife crossings. Our aim is to acquire a simple, non-invasive, cost-effective method to identify and quantify animals using wildlife crossing structures. If successful, the technique will enable demographic and movement data to be collected that are requisite for accurately assessing the conservation value of wildlife crossings. The development of this technique will significantly advance our understanding of wildlife crossings as conservation tools to enhance population viability.