DOI
https://doi.org/10.25772/0bw1-2e46
Defense Date
2021
Document Type
Directed Research Project
First Advisor
Dr. Chris Ehrhardt
Second Advisor
Dr. Susan Greenspoon
Third Advisor
Dr. William Eggleston
Abstract
“Touch” DNA is evidence that consists of epidermal cells deposited by handling objects and it is becoming more common in evidence samples processed in crime labs. Because “touch” DNA evidence samples may be low-template DNA and are often mixtures, there is a need to both nondestructively estimate the amount of DNA present prior to DNA purification and to identify and characterize mixtures prior to DNA typing. The purpose of this study was to test the use of FITC-labeled anti-testosterone antibodies as a potential tool to estimate the number of contributors in two-, three-, and four-person mixtures of epidermal skin cells, as well as estimate the quantity of DNA in single-source epidermal cells. DNA mixture profiles can be difficult to interpret and any information prior to profile generation may aid in developing the most informed interpretation of the data. Single-source and mixture samples bound to anti-testosterone antibodies were assessed for their fluorescent intensities using a Guava® easycyteTM flow cytometer. Only some of the two- (3/18) and three-person (1/9) mixture fluorescence histograms correctly estimated the number of contributors but none of the four-person (0/3) mixtures were successful. Statistical analysis of single-source and mixture histogram distribution demonstrated some capability of predicting the presence of a mixture but requires further testing. To predict DNA content, stained single-source samples were collected from the flow cytometer and subjected to DNA extraction and quantification. Plotting of median fluorescent intensity against the DNA quantity of single-source donors revealed no apparent correlation, though most DNA quantities were not reliably detected. To provide support to the mixture and DNA quantity studies, the specificity of the anti-testosterone antibody was assessed by comparing its fluorescent intensity to a FITC-labeled non-specific isotype control. The anti-testosterone antibody demonstrated a substantially greater fluorescent intensity indicative of greater antibody binding and was determined to be more specific than the isotype control. To optimize fluorescent intensity for future studies, single-source samples were stained with anti-testosterone and FITC-labeled anti-dihydrotestosterone antibodies together at various volumes [2.5, 5, and 10 uL]. The optimal combination of the two antibodies may be 2.5 uL anti-testosterone and 10 uL anti-dihydrotestosterone but requires more testing. This study demonstrates that anti-testosterone antibodies have the potential to be used in a nondestructive technique to estimate the number of contributors in a mixture of epidermal skin cells by specific staining but may not be able to estimate DNA quantity in single-source samples.
Rights
© The Author(s)
Is Part Of
VCU Master of Science in Forensic Science Directed Research Projects
Date of Submission
5-11-2021