Defense Date
2023
Document Type
Directed Research Project
First Advisor
Susan Greenspoon, PhD
Second Advisor
Christopher Ehrhardt, PhD
Third Advisor
William Eggleston, PhD
Abstract
In a forensic investigation, STR profiles generated from biological evidence can frequently be informative to identify an individual or provide an investigative lead. However, STR typing does not provide information to answer activity-level questions which can be highly important to a criminal case, such as the Time Since Deposition (TSD) of biological evidence. Determining the relative time in which a biological sample was left at a crime scene would provide additional information regarding the timeline of a crime and a person of interest’s relationship to the scene. One proposed method of estimating TSD is to use flow cytometry to analyze biological samples after hybridization with fluorescently labeled antibody probes. Flow cytometry is a non-destructive technique which uses visible light scatter and fluorescence parameters to measure physical and biological characteristics of individual cells. This study investigated the use of antibody hybridization and analysis by flow cytometry as a means to estimate TSD in saliva samples. Saliva samples were deposited and collected on a week and month basis over a 180-day time course, as well as on a daily basis over a 7-day time course. Saliva cells were hybridized with an antibody targeting cytokeratin, a structural protein found in epithelial cell types. Cytokeratin staining patterns and cellular autofluorescence were measured over time to determine if changes in fluorescent signal correlated to TSD. Autofluorescence profiles were also compared with DNA degradation levels, measured by a DNA quantitation-specific degradation index and STR typing to determine if a correlation existed between fluorescent signal changes and DNA quality.
Results showed a strong correlation between autofluorescence and TSD over the 180-day time course. Variation in fluorescence levels was also observed within cell populations and between contributors that increased with time, causing overlap in fluorescent signal at a day-to-day level. Cytokeratin antibody signal appeared to decrease after about 7 days, followed by a consistent increase from 7 to 60 days. A weak correlation was also observed between autofluorescent signal and DNA degradation levels (R2 ~ 0.09). These results show some promise in utilizing autofluorescent signal as a tool to estimate TSD, though further studies are needed to assess how variation in cellular fluorescence within and between individuals can affect the estimation of time since deposition. Autofluorescence signal changes may potentially be used to estimate DNA quality prior to any DNA analysis.
Rights
© The Author(s)
Is Part Of
VCU Master of Science in Forensic Science Directed Research Projects
Date of Submission
4-29-2023