Although penalties for violating Italy’s laws on driving under the influence of drugs (DUID) are very strict, no national protocol is enforced in order to assess possible offences in traffic accidents and routine traffic patrols. We describe two protocols that were applied to assess 298 cases of potentially impaired drivers out of 1653 stopped in Perugia, a town in Central Italy. One protocol was the D.O.S. carried out by means of oral fluid and urine and blood for confirmatory analysis. The other “SALIVA” uses only oral fluid collected during the road checkpoints. Psychoactive drugs were present in about 30% of the suspected drivers under both protocols, with cannabis being most often detected. The number of drivers who were sanctioned for DUID was significantly higher under the SALIVA protocol. Because non-standardized procedures showed critical issues, it is necessary to harmonize DUID national protocols. Toxicological screening and confirmatory tests, with standard analytic procedures, should be developed.