Shedding light on shedders.

All previous examinations of the shedder status of individuals have been based on conclusions inferred from the amount of DNA deposited by donors after they have held an object for a fixed period of time. In all interpretations of shedder status experiments have involved a range uncertainties, especially in regards to results arising from studies carried out in different laboratories. These apply to the efficiency of the swab collecting DNA from the item touched, the amount of DNA left on the swab after attempts to recover it, and the percentage loss of DNA during the lysis and extraction processes. No previous study has attempted to mitigate these uncertainties or verify how much of the DNA deposited was collected through swabbing, how much DNA present on the swab was recovered or how much DNA is lost during the extraction process. We present a study that accurately measures the deposition, collection and amplification of DNA deposited by a range of donors allowing for an accurate determination of the shedder status of individuals. Eleven donors were asked to wash their hands and then deposit a thumbprint onto glass slides by making pressure for 15 seconds 0, 15, 60 and 180 minutes after handwashing. Both left and right thumbs were used and all testing was performed in triplicate. Measurement of the quantity of cellular material deposited on the slides was carried out using DiamondTM Nucleic Acid Dye and fluorescence microscopy on each of 264 thumbprints. Fluorescence microscopy was then used to demonstrate that all the DNA present on the slides was recovered by the swabbing operations and then direct PCR, using the Identifiler™ Plus kit, was used to ensure that none of the DNA present on swabs was lost during DNA profiling. The combination of using a DNA binding dye and direct PCR allowed an accurate means of measuring the extent to which individuals exhibit different extents of shedding. This small study, 11 donors, showed that individuals fell into one of three distinct groups: heavy, intermediate, and light shedders, regardless of the hand used. Crown