Discovery of specific protein markers in multiple body fluids and their application in forensic science

Identification of multiple body fluids is crucial for the reconstruction and corroboration of crime event. However, for the body fluids with high component similarities, such as peripheral blood and menstrual blood, reliable distinguishing markers are still lacking. Furthermore, a comprehensive protein marker assay for multiple body fluids is urgently necessary for complex crime events. Herein, we established a highly specific and detectable method for discovering protein markers in peripheral blood, menstrual blood, saliva, semen and vaginal fluid through integrating in-depth discovery proteomics and a two-step targeted screening approach. Four menstrual blood markers with high endometrial specificities were identified for differentiation from peripheral blood and exhibited moderate protein concentrations for reproducible analysis with a protein quantitation CV value of 8.66%. Finally, a targeted discrimination method with 16 protein markers was established. We successfully identified 47 blind samples with 100% specificity and detection rate, sourced from five types of body fluids and presented on matrices such as cotton, tissues, slides or fluid. Overall, this work developed an effective method for discovering body fluid biomarkers, obtained specific protein markers to identify five kinds of body fluids and their targeted monitoring will show great significance for forensic science.