Differential extraction

Differential extraction (also known as differential lysis) refers to the process by which the DNA from two different types of cells can be extracted without mixing their contents. The most common application of this method is the extraction of DNA from vaginal epithelial cells and sperm cells from sexual assault cases in order to determine the DNA profiles of the victim and the perpetrator. Its success is based on the fact that sperm cells pack their DNA using protamines (rather than histones) which are held together by disulfide bonds. The protamines sequester DNA from spermatozoa, making it more resilient to DNA extraction than DNA from epithelial cells. Differential extraction (also known as differential lysis) refers to the process by which the DNA from two different types of cells can be extracted without mixing their contents. The most common application of this method is the extraction of DNA from vaginal epithelial cells and sperm cells from sexual assault cases in order to determine the DNA profiles of the victim and the perpetrator. Its success is based on the fact that sperm cells pack their DNA using protamines (rather than histones) which are held together by disulfide bonds. The protamines sequester DNA from spermatozoa, making it more resilient to DNA extraction than DNA from epithelial cells. After determining that sperm cells are present (typically through staining and light microscopy) in a vaginal/rectal sample, the subject's epithelial cells are lysed by a standard DNA extraction method, like a phenol/chloroform extraction and their DNA extracted through normal means. The epithelial DNA in solution is removed and saved, while the sperm cell's DNA precipitates with the attached protamines. Differential extraction uses a chemical called dithiothreitol (DTT) to disrupt the sulfur bonds in the protamines in order to release its DNA. Once the DNA is detached from the protamines, it is prone to standard DNA extraction methods. This creates two different DNA fractions from one sample, hopefully that of the victim and that of the perpetrator. However, the described method is difficult to carry out because it is both very labor intensive and time-consuming, leading to a build-up of untested rape kits. An estimated 500.000 rape kits alone in the US. Greenspoon et al. reported improvements in sample processing efficiency and throughput using robotic automation. However, associated costs for implementation in conjunction with low-throughput quantities of samples may be impractical for forensic laboratories and cannot be justified. Recently a multistep nucleic acid extraction procedure was introduced to provide DNA lysates of highest quality. A self-sealing membrane allows a stepwise release and separation of DNA from mixed specimens. Implemented in a spin-column system, it is ideally suitable for DNA extraction procedures involving differential extraction of forensic samples such as epithelium, saliva or blood vs. sperms. Simple and reliable extraction protocols for both, stained samples as well as gynecological swabs, respectively, overcome the often claimed difficulties in differential extraction (e.g. losing a sperm pellet through several washing steps).