Laser Microdissection Separation of Pure Spermatozoa Populations From Mixed Cell Samples for Forensic DNA Analysis

• NCJ Number: 217268
• Author(s): Christine T. Sanders; Emily K. Reisenbigler; Daniel A. Peterson
• Date Published: February 2007
• Length: 99 pages
• Grant Number(s): 2004-DN-BX-K215
• Agencies: NIJ-Sponsored
• Publication Type: Report (Study/Research)
• Award: Laser Microdissection Separation of Pure Spermatozoa Populations from Mixed Cell Samples for Forensic DNA Analysis

Annotation

This study assessed the feasibility of using laser microdissection (LMD) technology in order to achieve the precise separation of cell mixtures from both perpetrators and victims in sexual assault cases for subsequent DNA analysis.

Abstract

The study found that both hematoxylin/eosin and nuclear fast red staining performed well in differentiating sperm (from the perpetrator of sexual assault) and epithelial cells (from the victim) without compromising the subsequent procedures for DNA analysis. Both QIAamp and Lyse-N-Go methods were useful for recovering DNA from the sperm cells collected from LMD. The DNA extraction yields from cells collected with LMD ranged from 12-45 percent. LMD separation provided clear STR profiles of the male donor, separated from the epithelial cells from the victim. LMD outperformed preferential lysis in the separation of sperm from increasing proportions of epithelial cells. The study concludes that the method developed would be appropriate for use with the LMD collection of over 75 sperm cells when using typical forensic PCR conditions. In order to evaluate the separation capability of LMD, mixed cell samples obtained from semen and buccal swabs were examined. Collections of 75, 150, and 300 stained sperm cells were separated by LMD from mixtures, followed by STR analysis. First, the minimum number of LMD-collected cells necessary for STR typing was determined by using low-copy-number (LCN) protocols. Second, a comparative study was done on the LMD method and the preferential lysis method for separating sperm from increasing proportions of female epithelial cells for LCN analysis. Finally, four sexual assault case studies were examined with the use of LMD and LCN analysis. 17 figures, 5 tables, and 61 references