Abstract
Determination of skin wound vitality is an important issue in forensic practice. No reliable biomarker currently exists. Quantification of inflammatory cytokines in injured skin with MSD® technology is an innovative and promising approach. This preliminary study aims to develop a protocol for the preparation and the analysis of skin samples. Samples from ante mortem wounds, post mortem wounds, and intact skin ("control samples") were taken from corpses at the autopsy. After an optimization of the pre-analytical protocol had been performed in terms of skin homogeneisation and proteic extraction, the concentration of TNF-α was measured in each sample with the MSD® approach. Then five other cytokines of interest (IL-1β, IL-6, IL-10, IL-12p70 and IFN-γ) were simultaneously quantified with a MSD® multiplex assay. The optimal pre-analytical conditions consist in a proteic extraction from a 6 mm diameter skin sample, in a PBS buffer with triton 0,05%. Our results show the linearity and the reproductibility of the TNF-α quantification with MSD®, and an inter- and intra-individual variability of the concentrations of proteins. The MSD® multiplex assay is likely to detect differential skin concentrations for each cytokine of interest. This preliminary study was used to develop and optimize the pre-analytical and analytical conditions of the MSD® method using injured and healthy skin samples, for the purpose of looking for and identifying the cytokine, or the set of cytokines, that may be biomarkers of skin wound vitality.