The scientific community has responded to the misidentification of human cell lines with validated methods to authenticate these cells; however, there are few assays available for nonhuman cell line identification. We have developed a multiplex polymerase chain reaction (PCR) assay that targets tetranucleotide repeats in the mouse genome consisting of primers that amplify nine mouse short tandem repeat (STR) markers.
Unique profiles were obtained from fifty inbred mice and six mouse cell lines which were used to determine the allele distribution for each STR marker. Correlations between allele fragment length and repeat number were confirmed with sequencing. STR genotypes for L929 and NIH3T3 cell lines were shown to be stable with increasing passage numbers as there were no significant differences in fragment length in samples of low passage when compared to high passage samples. In order to detect cell line contaminants, primers for two human STR markers were incorporated into the multiplex assay to facilitate detection of human and African green monkey DNA. Adoption of this assay would increase confidence in studies using mouse cell lines and could ultimately save time and reduce costs by identifying misidentified or contaminated cell lines in advance.