The efficacy of Diamond™ nucleic acid dye-stained cell counting techniques for forensic application

Touch DNA is one of the most common types of biological material collected during criminal investigations. Diamond™ Nucleic Acid Dye (DD) has been shown to aid in touch sample visualisation and target sampling. It has also been used as a method of shedder categorisation that is cheaper and quicker than DNA methods. However, the DD method routinely involves manual cell counting, which can result in intra and inter-person variability similar to other manual techniques used in forensic science, for example, fingerprint identification. Additionally, DD based shedder categorisation involves counting cells in a portion of the touch deposit to extrapolate an individual's shedder status, and the sampling effect of such estimations is currently unknown. The present study tested different data analysis aspects of the DD method, including counting variability within and between people, shedder classification differences based on different counting methods (entire thumbprint, sub-section of a print with most cells, sub-section of a print deemed most representative of the entire thumbprint, and random sections), the use of ImageJ software to semi-automate counting and the use and extension of the DD method for investigating DNA Transfer, Persistence, Prevalence and Recovery (DNA-TPPR). The results of this study show that there are meaningful differences observed during counting processes both between and within people. These differences tended to increase as the factor of time, or the duration of counting, rather than the complexity of cell deposits being assessed. Investment in cell counting software that eliminates personal factors, such as boredom fatigue, can remedy most of these issues, however, will require optimisation, such as fibre recognition. Shedder testing was shown to be affected by the choice of sampling and categorisation methods, and suggested that using an entire finger or larger section size can provide increased precision. Finally, inverted worn gloves stained with DD may provide an acceptable alternative for hands in DNA-TPPR investigations, providing an interesting alternative for future research.