In this chapter we provide practical suggestions, together with examples, to identify, monitor and mitigate railway barrier effects on wildlife, as this is considered one of the railways’ greatest impacts. Railways can be both physical and behavioral barriers to wildlife movement, as well as disturbance to populations living close to them. Also, mortality is recognized as an important contribution to the barrier effect. However, the consequences of habitat loss, and fragmentation due to railways alone remain largely unexplored. Barrier effects have mainly been mitigated with wildlife passes, with the effectiveness of this tool being one of the most-studied topics in Railway Ecology. Methods formerly employed to monitor pass usage, such as track beds or video-surveillance, are now being replaced by molecular ones. Among the latter methods, genetic fingerprinting allows individual-based approaches, opening the door to population-scale studies. In fact, genetic sampling allows for the assessment of functional connectivity, which is closely linked to successful reproduction and population viability, variables not necessarily coupled with crossing rates. There is strong evidence that railway verges offer new habitats for generalist species and for opportunistic individuals, a point that deserves to be experimentally explored in order to find wildlife-friendly policies. Preventing animals from crossing (eg, by fencing), should be reserved for collision hotspots, as it increases barrier effects. Instead, it has been shown that warning signals or pole barriers effectively reduce collisions without increasing barrier effects. Palabras clave: Barrier, Railway