Micromanipulation of single cells and fingerprints for forensic identification.

Crime scene samples often include biological stains, handled items, or worn clothes and may contain cells from various donors. Applying routine sample collection methods by using a portion of a biological stain or swabbing the entire suspected touched area of the evidence followed by DNA extraction often leads to DNA mixtures. Some mixtures can be addressed with sophisticated interpretation protocols and probabilistic genotyping software resulting in DNA profiles of their contributors. However, many samples remain unresolved, providing no investigative information. Samples with many contributors are often the most challenging samples in forensic biology. Examples include gang rape situations or where the perpetrator's DNA is present in traces among the overwhelming amounts of the victim's DNA. If this is the only available evidence in a case, it is of paramount importance to generate usable information. An alternative approach, to address biological mixtures, could be the collection of individual cells directly from the evidence and testing them separately. This method could prevent cells from being inadvertently blended during the extraction process, thus resulting in DNA mixtures. In this study, multiple tools coupled with adhesive microcarriers to collect single cells were evaluated. These were tested on epithelial (buccal) and sperm cells, as well as on touched items. Single cells were successfully collected but fingerprints were swabbed in their entirety to account for the extracellular DNA of these samples and the poor DNA quality of shed skin flakes. Furthermore, micromanipulation devices, such as the P.A.L.M.® and the Axio Zoom.V16 operated manually or with a robotic arm aureka®, were compared for their effectiveness in collecting cells. The P.A.L.M.® was suitable for single cell isolation when smeared on membrane slides. Manual or robotic manipulations, by utilizing the Axio Zoom.V16, have wider applications as they can be used to isolate cells from various substrates such as glass or membrane slides, tapes, or directly from the evidence. Manipulations using the Axio Zoom.V16, either with the robotic arm aureka® or manually, generated similar outcomes which were significantly better than the outcomes by using the P.A.L.M.®. Robotic manipulations using the aureka® produced more consistent results, but operating the aureka® required training and often needed re-calibrations. This made the process of cell manipulations slower than when manually operated. Our preferred method was the manual manipulations as it was fast, cost effective, required little training, but relied on a steady hand of the technician.