A novel fluorescence-based method in forensic science for the detection of blood in situ

Full DNA profiles can be generated from just a few cells; however these profiles can be contaminated from other cell types present at the crime scene. We report here on the development of an immunofluorescent technique to spatially locate human-specific blood in situ and also on the ability of this technique to detect individual leukocytes and the DNA contained within them. Four monoclonal mouse anti-human antibodies were evaluated; anti-glycophorin A to detect erythrocytes and anti-CD45, anti-myeloperoxidase (MPO) and anti-histone H1 to detect the nucleated leukocytes. Each antibody was labeled with either Alexa Fluor 488 or 568 for direct application to blood smears which allowed the simultaneous detection of erythrocytes and leukocytes. Furthermore, because histones are DNA binding proteins, the application of anti-histone H1 allowed the detection of DNA within a blood smear. Importantly it was found that full DNA profiles could be achieved after using this method with similar peak area ratios compared to untreated cells. The fluorescent antibodies were found to be human-specific with the exception of anti-histone H1 due to its conserved sequence. However, used in combination with anti-CD45 or anti-MPO the location of DNA from human-specific leukocytes could be detected. The technique was also tested on older blood stains and was still found to be sensitive and cell-specific after 4 months. Following the optimization of the methodology, the fluorescent antibodies were applied to short lengths of black cotton fibres covered with human blood spots. Although the background fluorescence from the cotton was found to be high, erythrocytes and even individual leukocytes could easily be detected, indicating that this technique could be used to detect extremely minute amounts of blood. Used in combination with laser capture microdissection (LCM), this method could be used to pick off individual leukocytes for LCN DNA techniques.     

Analysis of hair after contamination with blood containing cocaine and blood containing benzoylecgonine

In post-mortem work, blood is a potential source of external contamination of hair. The present study was carried out to investigate the amount of drug absorbed into hair which has been contaminated with blood containing either cocaine or BE. Solutions were prepared containing 0.05, 0.1, 0.2, 0.5 and 3.0 μg/mL of either cocaine or BE in human blood. Samples of approximately 3.2 g of drug-free hair were contaminated by soaking in the blood solutions for 5 min. They were then removed and left at room temperature. Approximately 0.5 g of hair was collected from each of the blood soaked hair samples at 6 h, 1, 2, 4 and 7 days after contamination. As each hair sample was collected it was shampoo-washed to prevent further drug absorption. Hair samples were analysed in triplicate using a fully validated method described previously. EME and cocaethylene were also measured in order to find out if cocaine or BE was breaking down to these compounds. Both cocaine and BE were absorbed into hair in significant concentrations when the concentration in the blood was 0.5 μg/mL or greater; cocaine was more readily absorbed than BE. Cocaine broke down to EME (<LOQ) at 0.5 μg/mL and to EME (>LOQ) and BE (<LOQ) at 3.0 μg/mL. When the blood concentration of cocaine was 0.5 μg/mL or less, there was no evidence of it breaking down to form BE. From the samples soaked in blood containing BE, there was no evidence of the BE breaking down. The absorption of drug into hair did not increase as the contamination period increased from 6 h to 7 days.     

Development of a Nomenclature System for a Canine STR Multiplex Reagent Kit*†

Abstract: Despite the popularity of dogs in US households, canine DNA evidence remains largely untapped in forensic investigations partially because of the absence of well‐defined forensic short tandem repeats (STRs), lack of standardized and validated PCR protocols, STR reagent kits, and poorly developed nomenclature. A nomenclature system was established based on internationally recognized recommendations for human forensic STRs for a recently developed canine STR reagent kit. Representative alleles were sequenced from each of the 18 STRs and the sex‐typing marker included in the kit. This study also reflects on the impact of point mutations, insertions, and deletions within and outside the STR core repeat structures. An understanding of the STRs’ sequence and repeat structures will enable development of a robust and reliable allele nomenclature and improve the accuracy and precision of allele fragment sizing in canine forensic profiling. The expected allele sizes have been calculated, and their repeat stuctures defined based on sequence information.     

A test of the effectiveness of speaker verification for differentiating between identical twins

This paper presents investigations into the ability of speaker verification technology to discriminate between identical twins. It is shown that whilst, in general, the genetic and non-genetic characteristics of voice are both of value to speaker verification capabilities, it is the latter which is highly beneficial in the separation of the speech of identical twins. It is further demonstrated that through the use of unconstrained cohort normalisation as a complementary means for the exploitation of such voice characteristics, the verification reliability can be considerably enhanced for both identical twins and unrelated speakers. Experiments were conducted using a bespoke clean-speech database consisting of utterances from 49 identical twin pairs. The paper details the problem in speaker verification posed by identical twins, discusses the experimental investigations and provides an analysis of the results.