The description of materials and methods encompasses the plasmids used; the extraction of nucleic acids, amplification, and analysis of PCR products; the processing of genomic DNA's; and statistical analysis. The results reported addressed principle, linearity, and specificity of the assay; the persistent positive results of a negative run control as an indication of the presence of ubiquitous DNA; false positive results caused by human template DNA; statistical analysis of template DNA contamination; and the detection limit defined by template DNA contamination. Two sources that contributed to DNA background were identified. The first one was interpreted as DNA present in chemicals and on equipment, and the second was caused by operator handling. The amounts were equivalent to 2.5 and 8.9 pg/mL of sample, and the estimated frequencies of contamination were 65 percent and 35 percent, respectively, resulting in an effective limit of detection of 17.4 pg/mL. Below this level, named effective laboratory background, a result could not be considered as authentic. Knowledge of these parameters is important for laboratories that analyze minute amounts of human DNA by PCR for purposes such as quantification, typing, and sequencing. 5 figures and 17 references