Detection of Epithelial Cells in Dried Blood Stains by Reverse Transcriptase-Polymerase Chain Reaction

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Abstract
The identification of menstrual blood stains can be improved by detection of messenger ribonucleic acid (mRNA) specific for epithelial (endometrial) cells. RNA molecules, however, are believed to be unstable and require careful sample processing. In this study, we have investigated the extraction of RNA suitable for reverse transcriptase-polymerase chain reaction (RT-PCR) from dried blood stains stored for up to six months. With a modified RNA isolation protocol, it was possible to obtain RNA from dried blood stains with at least 5 × 102 leukocytes. In an additional experiment, we evaluated the RNA isolation from mixed stains composed of leukocytes and T47D cells, a breast cancer-derived cell line with epithelial origin. Detection of 102 T47D cells in a total number of 105 leukocytes was possible by amplification of cytokeratin 19 mRNA and progesterone receptor-mRNA specific for hormonally regulated epithelial cells. In both experiments amplification results were not dependent on storage time with similar data from one day to six months. Furthermore, it was possible to identify dried menstrual blood samples by showing the presence of mRNA specific for epihelial cells. These results demonstrate for the first time, that RNA suitable for RT-PCR, can be isolated from forensic specimens stored up to at least six months, and that a small number of epithelial (endometrial) cells can be identified in dried blood specimens. Using this method, it will be possible to identify the origin of small and partially degraded blood samples which can be especially useful in forensic evaluation of cases with sexual offense.