The development of multiplex STR systems for use in forensic DNA analysis has revolutionized forensic biology in the past 10 years, with widespread use in forensic DNA laboratories since the commercial availability of these systems. The Promega PowerPlex 16 STR allows increased efficiencies and discrimination power by robustly typing in a single reaction, the 13 CODIS loci, Amelogenin, as well as the low stutter, highly polymorphic pentanucleotide STR loci, Penta D and E (1). Because of its abilities, an internal validation of the Promega PowerPlex 16 DNA typing system was conducted in 2004 by the Center of Forensic Science in Toronto, Canada and Northern Regional Laboratories. The validation study found the Promega PowerPlex 16 system to be robust, reliable, highly discriminating and suitable for providing casework and reference sample genotypes for the U.S. National Databank system. It was seen to be sensitive, reliable, and discriminating when employed. In addition, it provides increased discrimination power from a single amplification. The PowerPlex 16 system is compatible with existing detection platforms, as well as those currently being validated by the CFS. This paper describes the validation study and presents and reviews the results, and evaluates the data to assist in the development of interpretation guidelines for forensic casework samples. Figures and references