Defense Date
2023
Document Type
Directed Research Project
First Advisor
Sarah Seashols-Williams
Second Advisor
Tracey Dawson Green
Third Advisor
Jospeh Reiner
Abstract
Evidence containing biological mixtures can make DNA interpretation more difficult due to ambiguous allele calls and artifacts that may be present within electropherograms. Separating out these cell mixtures can be advantageous to the analysts’ analysis methods for identifying STR profiles with higher accuracy. A potential cell separation method that could be implemented in the beginning of the forensic DNA workflow is optical trapping.
This research utilized a 5W ytterbium linearly polarized laser split into two optical traps, paired with a microfluidic device to isolate spermatozoa and vaginal epithelial cells from a 1:1 mixture of vaginal fluid and 1:20 diluted seminal fluid. Between 7 to 20 epithelial cells and 19 to 40 spermatozoa were isolated from the mixture and processed downstream to analysis. After DNA extraction the samples were quantified and determined to have yields higher than the theoretical amount of DNA present. Five of the samples had a degradation index above one, suggesting that degradation had occurred while the other four samples fell below one. A sample containing 21 trapped sperm cells had the highest degradation index of 15.5 and did not generate any STRs to be analyzed after capillary electrophoresis.
A complete single source profile was created with 40 trapped spermatozoa cells from the mixture. Two samples that contained 11 and 12 trapped epithelial cells had 94.87% of the expected alleles in the sample observed. However, there was drop-in present in the trapped epithelial cell samples. Single source profiles were generated from two samples containing epithelial cells and one sample containing sperm cells that were consistent with the vaginal fluid and seminal fluid donor respectively.
This research has shown a microfluidic device coupled with a dual trap laser for optical trapping can be an improved cell separation technique between spermatozoa and epithelial cells. This research also considered improvements with microfluidic flow control and dissociating clumped epithelial cells to increase the ability to readily trap targeted epithelial cells within a mixture. Nuclear dyes were tested in conjunction with this set-up to more easily identify cells that contain nuclear material. In summary, optical trapping may be an alternative method to separate out cells of interest prior to the DNA workflow.
Rights
© The Author(s)
Is Part Of
VCU Master of Science in Forensic Science Directed Research Projects
Date of Submission
5-5-2023