Y-chromosome short tandem repeats (Y-STR's) provide valuable information in cases of rape and questioned paternity, and they allow for the genetic identification of male lineages. An internal validation of the Y-STR multiplex was performed in accordance with the DNA Advisory Board Quality Assurance Standard. An internal validation verifies procedures that were established by another laboratory to ensure that they will be effective in one’s own laboratory. The internal validation used control and reference DNA samples. The validation studies included optimization of the PCR reaction, analysis of father-son pairs, development of population databases, and mixture and sensitivity studies. The Y-STR 10-plex examines 10 non-overlapping loci on the Y-chromosome. The 10-plex uses forward and reverse primer sets that reduce the lengths of the PCR amplicons, thereby making them more applicable in the analysis of the degraded samples encountered in forensic casework. The study demonstrated that the 10-plex is sensitive to 0.125 ng of input DNA and that female samples mixed with male samples did not interfere with Y-STR haplotyping. In a sample of 525 males, there were 3 instances of locus multiplication, 2 at DYS19, and 1 at DYS435. The results suggest that the Y-STR 10-plex is highly discriminating and can effectively distinguish male lineage. The Y-STR 10-plex would be an excellent addition to the autosomal STRs already employed in many forensic and paternity laboratories. 3 figures, 5 tables, 24 references