| The work presented in this dissertation is stitched together in a seamless manner by one common thread - the development of a microfluidic device technology for forensic DNA analysis. Our efforts have been focused on developing methods for the sample processing steps of forensic DNA profiling, specifically for analysis of sexual assault and blood evidence.With sexual assault evidence, conventional differential extraction is utilized to obtain separate male and female fractions of DNA, from which a profile from the victim and perpetrator can be delineated. Chapter 2 discusses the development of a microscale method to replace the conventional differential extraction based upon the different physicochemical properties of the two cell types. Chapter 3 focuses on the development of a quantitative PCR (gPCR) method for the simultaneous quantitation of male and total autosomal DNA in a biological sample as a means to evaluate the results of the cell sorting methods described here. Chapter 4 describes the development of a second means of obtaining separate male and female fractions of DNA from sexual assault evidence, this time exploiting ultrasonic energy to selectively trap cells in a flow stream in a microdevice. This 'acoustic trapping', in combination with simplistic, but effective, valving via precise flow control resulted in separation of the sperm cell and female DNA fractions.Chapter 5 describes the development of a capillary electrophoretic immunoassay (CE-IA) for the identification and species testing of suspected bloodstains. While not technically considered a processing step of DNA analysis, identification and species testing of suspected bloodstains is routinely conducted by forensic laboratories prior to DNA analysis. Upon translation to the microscale, this method would allow testing at the crime scene to help investigators best determine the evidence to be collected and sent to the forensic laboratory for DNA testing.The research described in this dissertation lays the foundation for future implementation of microdevices in forensic laboratories. The completion of these steps on the microscale is toward the development of fully-automated, integrated sample processing on microdevices, which could ultimately lead to rapid DNA analysis in forensic laboratories. |
