Hair fibres, a valuable form of biological evidence, are continuously shed, chemically stable, persistent, and easily transferred. Taxonomic classification of hair recovered during a criminal investigation provides crucial forensic intelligence and aids in evidence prioritisation. Traditional classification methods rely heavily on the analysis of hair shaft morphology, a practice that’s been heavily scrutinised in recent years. The rise of mass spectrometry-based proteomics provides a molecular alternative, leveraging genetic identifiers that exist in the proteome of each species in the form of amino acid sequence variants for taxonomic classification of biological material.
Here developed a proteomics workflow for the characterisation of taxonomically diagnostic peptides, isolated from digested telogen hair shafts of 15 species. Taxa included were selected with a specific focus on taxonomies of notable evidentiary value to forensic intelligence, namely humans and common domesticated species. A panel of 226 taxonomically informative peptides were identified and characterised, 209 which appear novel to this study. The biomarker panel includes 59 peptides with single-species specificity. Whilst, the remaining peptide biomarkers were diagnostic due to restricted taxonomic distribution, representing common or convergent evolution.
The panel of taxa-discriminating markers was subsequently integrated into a forensic workflow, optimised for universal sample preparation and standardised LC/MS/MS data acquisition. The developed workflow was used to analyze single fur hairs (20 mm), from three individuals from each species of interest. The data was processed using a curated protein database, providing an efficient and validated bioinformatics workflow that did not require prior source information.
The implementation of this panel of diagnostic peptides identified and characterised using a mass spectrometry-based proteomics workflow introduces a robust and novel biomolecular approach for forensic analysis of trace biological material.