Analysis of genetic variation in DNA sequences serves as a powerful method for human identification owing to its exceptional discriminative power for distinguishing individuals. In cases where DNA is compromised in recovered forensic evidence, other approaches are needed to achieve a similar level of differentiative potential for human identification. Proteins offer a promising alternative, particularly in recovered hair evidence where minimal intact genomic DNA remains, as hair proteins often persist for long periods of time and their amino acid sequences derive from DNA. Detection of amino acid polymorphisms in hair proteins as genetically variant peptides (GVPs) permits inference of individualizing single nucleotide polymorphisms for identification. Expanding upon previous proof-of-concept work, this research interrogates the human hair proteome to address fundamental questions about how experimental variables affect GVP detection success rates, and to bridge the gap between laboratory-optimized studies and application of this protein-based approach in routine forensic analysis. Effects of intrinsic variation to hair protein chemistry, including differences among body locations and with increasing hair age, and external exposures, such as an explosive blast, on variant peptide marker detection are investigated using trypsin digestion and liquid chromatography-tandem mass spectrometry (LC-MS/MS). This multi-disciplinary work demonstrates success in GVP detection and advances in knowledge of protein chemistry in hair as a function of different body locations, in aged hairs, and in damaged hairs recovered after an explosive blast, providing greater confidence in GVP analysis for forensic investigations. Not limited to forensic proteomics, these findings may be applicable to the wider bioanalytical sciences, including the medical, material, and agricultural sciences.
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