Improving body fluid identification in forensic trace evidence-construction of an immunochromatographic test array to rapidly detect up to five body fluids simultaneously.

Body fluid identification is a substantial part of forensic trace analyses. The correct determination of the origin of a biological stain may give valuable information regarding the circumstances of a crime. A simple way to detect a body fluid in a stain is the use of immunochromatographic strip tests. They are easy to use, user-independent, quick, and cheap. Currently, however, it is only possible to analyze one body fluid at a time, requiring the analyst to make previous, possibly subjective, assumptions on the body fluid at hand. Also, identification of mixed body fluids requires the use of several tests, which results in additional sample and time consumption. To combine a simple approach with the possibility to simultaneously detect several body fluids, we constructed a combined immunochromatographic strip test array based on commercially available tests. The array rapidly detects up to five body fluids with a single analysis, and allowing for subsequent DNA extraction from the same material. With this test it was possible to identify the components of a mixture, the test was easily incorporated into standard laboratory work, and its sensitivity and specificity were shown to be comparable to those of conventional strip tests.

Marker evaluation for differentiation of blood and menstrual fluid by methylation-sensitive SNaPshot analysis.

The differentiation of blood and menstrual fluid is especially important in cases of alleged sexual assault. While the identification of blood is relatively straightforward, the identification of menstrual fluid in trace evidence has been shown to be more challenging. This may be due to the complex nature of menstrual fluid that leads to intra- and inter-individual differences in composition. Nevertheless, recent advances in DNA methylation profiling have revealed promising markers for the differentiation of the two body fluids and furthermore, markers to distinguish menstrual fluid from vaginal fluid. A literature study was performed and in total, 11 markers were evaluated in this study of which seven could be validated for menstrual fluid and blood identification purposes. Marker "BLU2" (chr16:29757334) was identified as most suitable for differentiation of blood and menstrual fluid.

Forensic differentiation between peripheral and menstrual blood in cases of alleged sexual assault-validating an immunochromatographic multiplex assay for simultaneous detection of human hemoglobin and D-dimer.

Sexual assault is a serious offense and identification of body fluids originating from sexual activity has been a crucial aspect of forensic investigations for a long time. While reliable tests for the detection of semen and saliva have been successfully implemented into forensic laboratories, the detection of other body fluids, such as vaginal or menstrual fluid, is more challenging. Especially, the discrimination between peripheral and menstrual blood can be highly relevant for police investigations because it provides potential evidence regarding the issue of consent. We report the forensic validation of an immunochromatographic test that allows for such discrimination in forensic stains, the SERATEC PMB test, and its performance on real casework samples. The PMB test is a duplex test combining human hemoglobin and D-dimer detection and was developed for the identification of blood and menstrual fluid, both at the crime scene and in the laboratory. The results of this study showed that the duplex D-dimer/hemoglobin assay reliably detects the presence of human hemoglobin and identifies samples containing menstrual fluid by detecting the presence of D-dimers. The method distinguished between menstrual and peripheral blood in a swab from a historical artifact and in real casework samples of alleged sexual assaults. Results show that the development of the new duplex test is a substantial progress towards analyzing and interpreting evidence from sexual assault cases.

Sudden infant death syndrome: exposure to cigarette smoke leads to hypomethylation upstream of the growth factor independent 1 (GFI1) gene promoter.

PURPOSE: Smoking during pregnancy has long been known as an important risk factor for sudden infant death syndrome (SIDS). However, the precise relationship between the smoking behavior of the mother and SIDS still remains unclear. In this study, the influence of prenatal smoking exposure on the childrens' DNA methylation state of a CpG island located upstream of the promoter of the growth factor independent 1 (GFI1) gene was analyzed. METHODS: Blood samples of well-defined SIDS cases with non-smoking mothers (n = 11), SIDS cases with smoking mothers during pregnancy (n = 11), and non-SIDS cases (n = 6) were obtained from a previous study and methylation states were determined by bisulfite sequencing. RESULTS: Significant hypomethylation was observed in this CpG island in SIDS cases with cigarette smoke exposure compared to non-exposed cases. The strongest effect in this CpG island was observed for 49 CpG sites located within a transcription factor binding site. Coding for a transcriptional repressor, GFI1 plays an important role in various developmental processes. Alterations in the GFI1 expression might be linked to various conditions that are known to be associated with SIDS, such as dysregulated hematopoiesis and excessive inflammatory response. CONCLUSION: Data obtained in this study show that analysis of methylation states in cases of sudden infant death syndrome might provide a further important piece of knowledge toward understanding SIDS, and should be investigated in further studies.

Validation of an immunochromatographic D-dimer test to presumptively identify menstrual fluid in forensic exhibits.

Identifying the biological source of a crime scene stain can be crucial for police investigations in many scenarios. Blood is one of the most common fluids found, and accurate differentiation between peripheral blood and menstrual fluid could provide valuable information regarding the issue of consent in sexual assault cases. For the detection of menstrual fluid, no easy-to-use presumptive test is available to date. Therefore, this study aimed to validate a simple immunochromatographic test for the indication of menstrual fluid, focusing on a D-dimer assay. The Clearview® rapid D-dimer test provides a diagnostic assay for the detection of fibrin degradation products. We validated the sensitivity and robustness of the assay using fresh and dried menstrual fluid samples, body fluid mixtures, diluted samples, and casework swabs. Cross reactivity was tested for saliva, semen, vaginal fluid, and blood. No false positive results were obtained; it was possible to successfully analyze mixtures, highly diluted samples, and casework swabs. The results of this study indicate that the D-dimer assay reliably detects menstrual fluid in forensic exhibits and is easy to implement into the current workflow of body fluid identification.

Independent validation of body fluid-specific CpG markers and construction of a robust multiplex assay.

Potential forensic use of tissue-specific DNA methylation markers has recently been discussed for the identification of the biological source of a stain. In this study 13 promising markers were evaluated to identify suitable candidate markers for the development of a robust and reliable multiplex assay. The results of this study suggest that a combination of only four highly informative markers will be enough for clear body fluid identification. A multiplex assay was developed for the identification of menstrual blood, saliva, semen, and venous blood. This assay was successfully applied to the identification of these body fluids in mixtures and crime scene stains. The multiplex assay aids in the identification of not only single source body fluids but also of body fluid mixtures. The main advantage of using DNA methylation assays over alternative tests is that it can be applied at a later time point in the investigative process since testing is possible even after DNA analysis.

Effect of systematic dental shape modification in bitemarks.

Studies on human cadaver models have reported significant levels of distortion of bitemarks in skin, indicating that tooth characteristics are not reliably transferred and recorded in the bitten subject. Moreover, matches among the anterior biting dentition in open population studies have been found. This prompts the question as to what degree of difference in shape will distinguish one dentition from another as reflected in a bitemark. In order to understand how these variables appear on skin, 10 dental casts with systematic variations in tooth positions were produced. The height of the lateral incisors was systematically altered in 1mm increments up to 3mm and lateral incisor/canines were altered in facial/lingual displacement in 1mm increments up to 5mm. Each of the models was used to produce a series of 10 repeated bites, distributed over arms and legs of un-embalmed cadavers. Landmark-based geometric morphometrics were used for analysis of digital images of the bitemarks. Results indicate that alterations of height and displacement of particular teeth affected the position of impressions created by the adjacent teeth. Displacement of one lateral incisor/canine led to a relative shift in impressions of the central incisors and unaltered canines, while height alteration of the lateral incisors led to a shift in relative position of central incisors as recorded in the bitemark. The prominence of displacements was more pronounced in the bitemarks than in images of the dentition used to make the bites, thus the bitemarks tended to exaggerate the differences. It was found that a displacement of 5mm between teeth allowed for reliable distinction between bitemarks. No such threshold of distinction could be established for differences in height of teeth under these experimental conditions. The effect of distortion was more significant in the mandibular than maxillary arch, suggesting that the mandible exhibits higher variation than the maxilla, as impressed in skin. Numerous bitemarks also exhibited arch flattening, consistent with recent studies showing arch width as the principal variable in a bitemark.