Sex-Based Targeted Recovery of Cells in a Heterogeneous Mixture: Separating Male and Female Like Cells

Mixture interpretation remains a central challenge in the forensic DNA field. The majority of solutions seek to ease the complexity mixture interpretation on the "backend", rather than address the separation of the individual biological components (e.g. cells). With individual components (contributors) of mixtures separated, the resulting profiles are simple to interpret (single-source), require less time and resources, and ultimately lead to increased confidence in the conclusions. Currently, there are no methods in use that allow sex-based separation of like-cell types, such as the epithelial cell mixtures of males and females. These scenarios are not uncommon in casework, and although Y-STR profiles may be generated, these profiles are not eligible for upload to DNA databases and, therefore, cannot be used effectively when the crime involves unknown perpetrators. This project proposes the development and optimization of a method to identify and profile male-specific cells recovered from a mixture of male and female-like cells. This 15-month project consists of three phases: (1) evaluation/optimization of a Y-chromosome labeling method for epithelial cells, (2) valuation/optimization on the DEPArray using pristine single-source and mixture samples, and (3) degraded single-source and mixture samples. The proposed method is a union of two accepted and well tested techniques, FISH-based immunohistochemistry for Y-chromosome labeling and DEPArray-mediated cell sorting/recovery. Benefits include enhanced resolution of the components of likecell mixtures of males and females, improved efficiencies and simple profile interpretation, fewer resources expended (time/money), and more probative investigative leads through generating CODIS eligible profiles. Note: This project contains a research and/or development component, as defined in applicable law, and complies with Part 200 Uniform Requirements - 2 CFR 200.210(a)(14). CA/NCF