Marco Scarpetta, Ph.D., Susan Greenspoon, Ph.D., Melanie Drayton, B.S. and Stefanie Turck, B.S.
Detroit Police Department, Forensic Services Division, Detroit, MI

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The Detroit Police Crime Lab has historically used Chelex as a method to isolate DNA for amplification and typing of bloodstains at the HLADQA1 and PM loci. Preliminary validation of several STR systems for casework has demonstrated that Chelex DNA is not suitable for STR amplifications for our purposes. Therefore, we have begun to extract DNA by the QIAamp spin column procedure as an alternative for amplification and typing of various STR loci. Additionally, we determined if QIAamp isolated DNA is suitable for DQA1, PM and D1S80 typing. Three studies were designed to address these questions.

The first study examined DNA isolated by the QIAamp versus the Chelex procedures on bloodstains from our database. This comparison study demonstrated that for routine bloodstains, QIAamp DNA is, in fact, a better substrate for amplification than Chelex. Next, a matrix study was performed to determine if the QIAamp DNA procedure would give better results on bloodstains deposited on "problem surfaces" such as leather, dirt and various dyed fabrics. Again, QIAamp DNA was more successfully typed than Chelex DNA. On several surfaces, QIAamp DNA was easily typed, while Chelex DNA gave little or no signal. Dirt was the only substrate on which Chelex DNA gave a weak, uninterpretable signal, while QIAamp DNA showed no signal at all.

Finally, a comparison study of three different methods including QIAamp spin columns, Centricon microconcentrators and ethanol precipitation was performed on nonprobative sexual assault casework samples. The non-sperm cell fraction was lysed according to the Perkin-Elmer protocol. The sperm cell fraction was incubated overnight in the presence of DTT for complete cell lysis. The cell fractions were split in triplicate, and the DNA was isolated from the non-sperm and sperm fractions of each sample by QIAamp, Centricon, and ethanol precipitation. Once the DNA extractions were completed, DNA concentrations were determined using the QuantiBlot kit (Perkin-Elmer) and total yields were calculated. Equal quantities of DNA from all samples were used in the amplification reactions. DNA isolated by the QIAamp procedure (approx. 30 min. after cell digestion) saved a considerable amount of time over the Centricon (65 min.) or ethanol (80min.) procedures. QIAamp DNA was also present at the greatest yield, with no decrease in the quality of the DNA or its ability to be reliably PCR amplified and typed using commonly employed methods. Therefore, we intend to validate the use of QIAamp spin columns for DNA isolation in casework.