Seratec (Germany)

Body fluid identification at crime scenes is the first step in the forensic biology workflow, leading to the identification of the perpetrator and/or, in some cases, the victim. Current methods that are regularly used in forensic criminal evidence analysis utilize well-studied properties of each fluid as the foundation of the protocol. Among these approaches, alternative light sources, chemical reactions, lateral flow immunochromatographic tests, and microscopic detection stand out to identify the main body fluids encountered at crime scenes: blood, semen, and saliva. However, these often come with limits for specificity and sensitivity. There is also difficulty with fluid mixtures, environmental degradation, and destruction of the sample by the method used. Other fluids, like vaginal fluid and fecal matter, lack standardized protocols and require innovative ideas for accurate analysis without compromising the sample. Emerging technologies based on molecular methods have been the focus of body fluid research, with emphasis on topics such as mRNA, microRNA, epigenetics, and microbial analysis. Additional information alongside the determination of fluid origin could be an advantage from new molecular techniques, such as the identification of donors from SNP analysis, if regular STR analysis is not possible. Validation studies and the integration of such research have the potential to expand and enhance the laboratory practices of forensic science. This article will provide an overview of the current methods applied in the crime lab for body fluid identification before exploring active research in this field, pointing out the potential of these techniques for application in forensic cases to overcome present issues and expand the variety of body fluids identified.

At crime scenes, apart from the detection of blood, it may be important to determine whether a person was alive at the time of blood deposition. Based on the rapid onset of fibrinolysis after death, this pathway could be considered to identify potential biomarkers for postmortem blood. Fibrinolysis is the natural process that breaks down blood clots after healing a vascular injury. One of its products, D-dimer, could be a potential biomarker for postmortem blood. SERATEC® (SERATEC® GmbH, Göttingen, Germany) has developed the PMB immunochromatographic assay to simultaneously detect human hemoglobin and D-dimer. The main goals of this study were to assess the possibility of using this test to detect postmortem blood, evaluate D-dimer levels in antemortem, menstrual, and postmortem blood, and assess the ability to obtain STR profiles from postmortem blood. Except for one degraded sample, all postmortem blood samples reacted positively for the presence of D-dimer using the SERATEC® PMB test. All antemortem blood samples from living individuals showed negative results for D-dimer detection, except for one liquid sample with a weak positive result, probably due to pre-existing health conditions. Menstrual blood samples gave variable results for D-dimer. The DIMERTEST® Latex assay was used for semi-quantitative measurement of D-dimer concentrations, with postmortem and menstrual blood yielding higher D-dimer concentrations compared to antemortem blood. Full STR profiles were developed for all postmortem samples tested except for one degraded sample, pointing to the possibility of not only detecting postmortem blood at the crime scene but also the potential identification of the victim.

This study re-introduces a protein-free rapid test method for nucleic acids on paper based lateral flow assays utilizing special multichannel nitrocellulose membranes and DNA-Gold conjugates, achieving significantly enhanced sensitivity, easier protocols, reduced time of detection, reduced costs of production and advanced multiplexing possibilities. A protein-free nucleic acid-based lateral flow assay (NALFA) with a limit of detection of 1 pmol of DNA is shown for the first time. The total production duration of such an assay was successfully reduced from the currently known several days to just a few hours. The simplification and acceleration of the protocol make the method more accessible and practical for various applications. The developed method supports multiplexing, enabling the simultaneous detection of up to six DNA targets. This multiplexing capability is a significant improvement over traditional line tests and offers more comprehensive diagnostic potential in a single assay. The approach significantly reduces the run time compared to traditional line tests, which enhances the efficiency of diagnostic procedures. The protein-free aspect of this assay minimizes the prevalent complications of cross-reactivity in immunoassays especially in cases of multiplexing. It is also demonstrated that the NALFA developed in this study is amplification-free and hence does not rely on specialized technicians, nor does it involve labour-intensive steps like DNA extraction and PCR processes. Overall, this study presents a robust, efficient, and highly sensitive platform for DNA or RNA detection, addressing several limitations of current methods documented in the literature. The advancements in sensitivity, cost reduction, production time, and multiplexing capabilities mark a substantial improvement, holding great potential for various applications in diagnostics, forensics, and molecular biology.

The main aim of this work was to validate the detection of saliva samples from denim, cotton, and polyester fabrics aged up to six months, applying rapid immunochromatographic tests resulting in the analysis of nuclear and mitochondrial DNA recovered. A comparison was also carried out between two saliva detection tests, Laboratory and Crime Scene. 50 μl saliva samples (three per time and test) were deposited on denim, cotton, and polyester fabrics. After 1, 3, 7, 14, 21, 30, 60, 90, 150, and 180 days of storage at room temperature, the samples were recovered by swabbing and detected by SERATEC® Amylase (Laboratory) test and SERATEC® SALIVA CS (Crime Scene) test (SERATEC®, Göttingen, Germany). DNA was isolated from the swab extraction buffer applying a silica-based methodology, and quantified based on fluorescent and human-specific quantifications. Then, it was submitted to STR profiling and mtDNA sequencing. According to our results, saliva stains up to six months after deposition remain valid specimens. The intensity of the bands varied among fabric type and time. Total DNA was successfully recovered from all tested samples, though with the limitations of obtaining partial nuclear DNA profiles from the oldest samples. In contrast, complete characterization of mtDNA was achieved from all samples. Lab and CS tests performed similar on the detection of saliva, as well as, DNA yield and profiling. Future research will be able to expand these results, analyzing the stability of other body fluids and the sensitivity of rapid immunochromatographic tests to detect them.