Expedited, chemically enhanced sperm cell recovery from cotton swabs for rape kit analysis

Autor: Jessica V. Norris, Sarah J. Linke, James P. Landers, Kate Manning, Jerome P. Ferrance
Rok vydání: 2007
Předmět:
Zdroj: Journal of forensic sciences. 52(4)
ISSN: 0022-1198
Popis: This report focuses on the development of a method for chemically induced enhancement of cell elution and recovery from cotton swabs. The method exploits the exclusive use of detergents for intact cell removal, and can be utilized in conjunction with, or to circumvent, conven- tional differential extraction (DE). Samples treated with Sarkosyl (54.4 € 1.8%) and sodium dodecyl sulfate (SDS) (78.5 € 0.7%) yielded higher sperm cell recoveries than a conventional DE buffer (39.4 € 2.1%). The results indicated that the choice of detergent affected sperm cell yield, with anionic detergents having the greatest effect. Storage time of samples affected the concentration of detergent required for optimal sperm cell recovery, longer times requiring increased detergent concentrations. In addition, the extent of sperm cell lysis by proteinase K digestion was evaluated. The results indicate that the exclusive use of SDS enhances the release of sperm and epithelial cells from a cotton swab as compared with DE buffer, providing for a more effective DNA analysis. Genetic analysis of perpetrator and victim DNA from vaginal cotton swabs is a well-established forensic technique for investi- gating and prosecuting sexual crimes (1-3). Analytical results suit- able for prosecution rely on separation of DNA from the perpetrator and victim to obtain individual DNA profiles. The cur- rent protocol for recovery of biological materials from a cotton swab involves differential extraction (DE), a method that utilizes proteinase K and an anionic detergent to selectively lyse vaginal epithelial cells while eluting sperm cells intact (2). Sperm cells are pelleted by centrifugation, and the supernatant containing the epi- thelial cell DNA is removed. The sperm cells are then resuspended in a buffer containing dithiothreitol (DTT), reducing the disulfide bond network in the sperm cell head and allowing the nuclear membranes to be lysed, releasing the male DNA. Differential extraction allows for independent recovery of male and female DNA from vaginal swabs containing mixtures of vagi- nal cells and spermatozoa, and has been routinely used in forensic laboratories since its introduction. However, sexual assault samples often contain sperm cells as a minor component in the presence of excess epithelial cells. DE is frequently ineffective for these sam- ples, as a considerable number of vaginal epithelial cells can remain undigested in the sperm cell fraction after the initial lysis step, resulting in an unfavorable ratio of male to female DNA (4). The probability of obtaining an interpretable male DNA profile is reduced when excess female DNA is present, as polymerase chain reaction (PCR) results may be obscured by excess female DNA
Databáze: OpenAIRE
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