Discovery of highly specific protein markers for the identification of biological stains

Abstract

DNA profiling has transformed the field of forensic biology by making it possible to individualize biological stains. The identification of the stain itself, however, continues to present forensic serologists with significant challenges. Current antibody- and enzyme activity-based assays yield only presumptive results as detection in nontarget body fluids or cross-reactivity with nonhuman sources have both been well documented. For other critical body fluids such as vaginal and menstrual fluids, there are no commercial tests at all. Using a three-pronged, comparative proteomic strategy based on proteome fractionation by HPLC followed by MS, a panel of 29 candidate protein biomarkers have been proposed as highly specific indicators of human saliva, urine, seminal fluid, vaginal fluid, peripheral blood, and menstrual fluid. The combination of consistent identification by multiple strategies in the current study; confirmation in independently compiled proteomic databases; and information on tissue expression and/or functionality from the proteomic literature all support the proposition that these proteins will have utility as reliable biomarkers of their target body fluids. The identification of candidate high-specificity protein biomarkers for human body fluids encountered in forensic investigations lays the foundation for the development of faster and more reliable approaches to the serological analysis of evidentiary stains.

High‐throughput miRNA sequencing and identification of biomarkers for forensically relevant biological fluids

“…The proteome consists of the proteins found in a particular tissue. For forensic body fluid identification, proteins uniquely found in a particular biological fluid could be exploited, but significant complications in the identification of proteins hamper this approach . Proteins undergo extensive PTM and tend to be low in abundance in fluids such as urine .…”

Verification of protein biomarker specificity for the identification of biological stains by quadrupole time‐of‐flight mass spectrometry

“…A comparative analysis of individual body fluids was previously conducted to assess the potential utility of proteins as biomarkers for the reliable identification of human body fluids. This approach resulted in the identification of five panels of candidate protein biomarkers that appeared to be highly‐specific for saliva, seminal fluid, vaginal/menstrual fluids, peripheral blood and urine . The apparent specificity of these candidate biomarkers, however, was based on the proteomes of just five individual humans per bodily fluid.…”

“…Comparative “whole‐proteome” consensus mapping using 1D and 2D HPLC in combination with electrospray ionization ion trap mass spectrometry had previously been used to identify protein biomarkers with potential utility for the identification of human body fluids . To more rigorously evaluate the specificity of these candidate protein biomarkers for their intended body fluid, a targeted Q‐TOF MS/MS platform was employed.…”

“…Currently, body fluid identification assays based on messenger RNA transcripts , microRNA expression profiles , differential epigenetic modifications , Raman spectroscopy patterns , and protein‐biomarker detection by mass‐spectrometry have garnered the most attention. Each of these proposed methods have their own strengths and weaknesses.…”

“…Mass spectrometry provides the ability to multiplex analyses with improved accuracy and specificity without the added costs or time required for antibody generation and characterization . The current study describes a biomarker verification workflow based on the use of a quadrupole time‐of‐flight (Q‐TOF) mass spectrometer to verify the specificity of 23 previously described candidate protein biomarkers for five target body fluids with forensic utility as well as the consistency of their expression among individuals in the general population.…”