The replication error phenotype, revealed by the observation of widespread microsatellite instability (MIN), has been identified as a new mechanism of cancer susceptibility, and the comparison of the allele sizes of polymorphic microsatellite repeats between normal and tumor DNA is now frequently undertaken in colorectal and other human neoplasias. The lack of precise characterization of the electrophoretic profiles of microsatellites is one of the main sources of discord between the rate of MIN reported for the same type of tumor by different investigators. The recent introduction of fluorescent-based semiautomated microsatellite analysis allows a more accurate size comparison, but one or more artificial peaks, generated during polymerase chain reaction (PCR) and/or electrophoresis, are frequently detected along with the true allele peaks. The aim of this study was to characterize the most frequent artificial extra peaks in the short tandem repeats (STRs) used by us to assess MIN in human cancers. We analyzed eight microsatellite loci in 113 primary brain tumors. HumFibra/FGA exhibited the most frequent extra peak formation. For each microsatellite there is a characteristic pattern of artifact formation which must be recognized to avoid a false-positive diagnosis of MIN.