Differentiation of five body fluids from forensic samples by expression analysis of four microRNAs using quantitative PCR

•Differentiation of five body fluids via analysis of four miRNAs.•Application of a validated, state-of-the-art quantitative PCR procedure.•Detection of blood in 36-year-old forensic samples.

Applying molecular genetic approaches for the identification of forensically relevant body fluids, which often yield crucial information for the reconstruction of a potential crime, is a current topic of forensic research. Due to their body fluid specific expression patterns and stability against degradation, microRNAs (miRNA) emerged as a promising molecular species, with a range of candidate markers published. The analysis of miRNA via quantitative Real-Time PCR, however, should be based on a relevant strategy of normalization of non-biological variances to deliver reliable and biologically meaningful results. The herein presented work is the as yet most comprehensive study of forensic body fluid identification via miRNA expression analysis based on a thoroughly validated qPCR procedure and unbiased statistical decision making to identify single source samples.