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SpermFACS: Validation of a Highly Sensitive Sperm Cell Sorting Method for Sexual Assault Casework Analysis.

Kristina Fokias1,2,3, Iene Rutten2,3, Karen Ven2,3

  • 1KU Leuven, Forensic Biomedical Sciences, Department of Imaging & Pathology, Leuven 3000, Belgium.

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|March 24, 2026
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Summary

A new fluorescence-activated cell sorting (FACS) method, spermFACS, efficiently separates sperm cells for forensic DNA analysis. This technique improves DNA profiling from challenging samples, aiding sexual assault investigations.

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Area of Science:

  • Forensic Science
  • Molecular Biology
  • Genetics

Background:

  • Sperm cell identification is crucial in sexual offense investigations.
  • Differential lysis (DL) is the standard but often inadequate method for sperm separation.
  • Limitations of DL include time consumption and poor DNA profiles from low sperm-to-epithelial cell ratios.

Purpose of the Study:

  • To establish and validate a fluorescence-activated cell sorting (FACS) method, termed spermFACS, for routine sperm cell separation from vaginal swabs.
  • To meet ISO/IEC 17025 guidelines for forensic implementation.
  • To assess spermFACS's effectiveness in challenging forensic samples.

Main Methods:

  • Development and validation of a FACS-based methodology (spermFACS).
  • Separation of sperm cells from vaginal swab samples.
  • Comparative analysis of spermFACS against differential lysis (DL).
  • Assessment of DNA profile quality from mixed cell populations and aged samples.

Main Results:

  • spermFACS successfully recovered informative male autosomal DNA profiles from mixtures with sperm/epithelial cell ratios as low as 1:7500.
  • Effective DNA profiling was achieved from postcoital samples collected up to 120 hours after intercourse.
  • spermFACS demonstrated superior separation efficiency and purity compared to DL, supported by likelihood ratio calculations.

Conclusions:

  • spermFACS is a valuable tool for forensic analysis, particularly in delayed-report sexual assault cases.
  • The method shows potential for deconvoluting complex mixtures, such as in multiple offender rapes.
  • Integrating spermFACS into forensic workflows can enhance offender identification and case resolution.