Multiplex short tandem repeat (STR) analysis has emerged as the dominant forensic DNA identification method because it is easy to interpret, can use sub-nanogram amounts of DNA, has a high degree of discrimination, and can yield results in 1-3 hours. In the United States, these advantages have led to development of a national felon database employing 13 core STR loci. In May 2000, Promega Corporation introduced the PowerPlex 16 as the first multiplex system capable of simultaneously amplifying all 13 core STR, the sex determinant locus, amelogenin, and two high discrimination low stutter pentanucleotide STR loci, Penta D and Penta E. This study presents validation data from 24 laboratories and developmental data from Promega Corporation. These data demonstrate the robustness of the system and the ability to successfully use the PowerPlex 16 System with casework samples in a large number of forensic laboratories. Tables, figures, references