Bone Proteomics Method Optimization for Forensic Investigations

Area of Science:

• Forensic proteomics
• Biomolecular analysis
• Mass spectrometry

Background:

• Proteomic analysis of skeletal remains aids forensic investigations.
• Maximizing protein recovery and minimizing laboratory errors are crucial for forensic casework.
• Standardization of proteomic workflows is needed for reliable results.

Purpose of the Study:

• To compare different bone protein extraction and analysis workflows for forensic applications.
• To evaluate the impact of novel S-Trap technology and data acquisition methods (DDA vs. DIA) on proteomic outcomes.
• To enhance the reproducibility and sensitivity of proteomic analysis in forensic bone samples.

Main Methods:

• Comparison of in-StageTip, S-Trap with lysis solution 1, and S-Trap with lysis solution 2 workflows.
• Analysis of 30 human cortical tibiae samples using liquid chromatography with tandem mass spectrometry (LC-MS/MS).
• Evaluation of data-dependent acquisition (DDA) versus data-independent acquisition (DIA) for protein identification.

Main Results:

• S-Trap workflows yielded higher proteome recovery and reduced induced deamidations compared to in-StageTip.
• Data-independent acquisition (DIA) demonstrated greater sensitivity and broader protein identification, especially for low-abundance proteins.
• Open-source software enhanced protein identification in both DDA and DIA modes.

Conclusions:

• The S-Trap protocol is well-suited for forensic bone proteomic analysis.
• Combining S-Trap with DIA offers improved proteomic outcomes and reproducibility for forensic applications.
• This optimized workflow contributes to the standardization of bone proteomic analyses in forensic science.