Identification evidence unraveled by strict proteomics rules toward forensic samples

Area of Science:

• Proteomics
• Forensic Science
• Toxicology

Background:

• Current snake venom identification relies on morphology and immunoassays, lacking precision.
• Proteomics offers a powerful alternative for detailed venom analysis.
• Forensic applications require highly accurate and validated identification methods.

Purpose of the Study:

• To apply bottom-up proteomics for identifying proteins and species in suspected snake venom samples.
• To develop and validate strict identification criteria for forensic applications.
• To assess the toxicity of identified venom components.

Main Methods:

• Ultrahigh-performance liquid chromatography-quadrupole-electrostatic field Orbitrap tandem mass spectrometry (UHPLC-QE-Orbitrap-MS/MS) for protein identification.
• In-sol and in-gel digestion for sample pretreatment.
• Nontargeted and targeted data acquisition modes for comprehensive analysis.
• Database matching against SWISS-PROT and a specific taxonomy database.
• Cytotoxicity assays to determine venom toxicity.

Main Results:

• Successfully identified proteins and species attribution in seized snake venom samples.
• Established strict protein identification rules: high confidence (FDR < 1%), ≥2 unique peptides, and <5 ppm mass accuracy for unique peptides.
• Identified samples as cobra venom, containing 10 abundant proteins, including acid phospholipase A2 and three-finger toxins.
• Determined cytotoxins to be the most abundant and likely most toxic proteins.

Conclusions:

• Proteomics provides a robust method for accurate snake venom identification in forensic contexts.
• The developed strict identification criteria meet forensic science standards for reliability.
• This approach enhances the capability to identify venom sources and understand their toxicological profiles.