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P118 – Towards selective male DNA profiling: A microfluidic strategy for epithelial cell mixtures
More sexual assault cases are reported globally, with 1 in 3 women experienced physical and/or sexual violence in their lifetime. In this context, forensic investigations aim to identify the perpetrator through the analysis of DNA evidence left behind during the crime. When ejaculation does not occur or sperm is insufficient, epithelial cell-derived traces represent an alternative source of cellular material from the perpetrator. However, such samples typically contain few cells with high-quality DNA or a mixture of cells from multiple contributors, which limits the applicability of standard technologies.
Here, we present a microfluidic-based strategy for the selective isolation and enrichment of male cells from male-female cell mixtures, with the aim of improving the analysis of mixed DNA epithelial trace evidence and enabling the generation of more accurate and informative DNA profiles .
Our approach employs a commercially available polysaccharide-based semi-permeable capsule (SPC) formulation, leveraging droplet microfluidics for uniform, high-throughput droplet generation and single-cell encapsulation, followed by shell crosslinking and transfer into an aqueous suspension. By exploiting the intrinsic porosity of the capsules, individual nucleated male or female cells encapsulated within SPCs are lysed and subjected to in-SPCs male-specific PCR to selectively amplify male DNA sequences. PCR performance is qualitatively assessed based on the fluorescence signal of SPCs containing male-specific amplicons.
The approach is first established using separate on-chip systems for single source non-mixed male and female cell lines, while optimization for mixed samples is ongoing. In parallel, fluorescence-activated droplet sorting (FADS) is being evaluated for selective enrichment of male-derived SPCs based on the fluorescence signal generated from amplified male DNA sequences. Sorted SPCs are subsequently pooled and chemically broken to release their contents, and male enrichment in mixed samples is verified through gel electrophoresis by comparing the amplicon lengths of male control sample and enriched male sample.
This platform introduces a novel route for the identification of male perpetrators and contributors in sexual assault cases involving female victims based solely on epithelial trace DNA.
Authors
- Bram Bekaert (KU Leuven/UZ Leuven, Belgium)
- Alice Rettore (KU Leuven, Belgium)
- Karen Ven (KU Leuven, Belgium)
- Iene Rutten (KU Leuven, Belgium)
- Jeroen Lammertyn (KU Leuven, Belgium)
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