Frank Heidorn

Single Cells for Forensic DNA Analysis—From Evidence Material to Test Tube

Abstract:  The purpose of this project was to develop a method that, while providing morphological quality control, allows single cells to be obtained from the surfaces of various evidence materials and be made available for DNA analysis in cases where only small amounts of cell material are present or where only mixed traces are found. With the SteREO Lumar.V12 stereomicroscope and UV unit from Zeiss, it was possible to detect and assess single epithelial cells on the surfaces of various objects (e.g., glass, plastic, metal). A digitally operated micromanipulator developed by aura optik was used to lift a single cell from the surface of evidence material and to transfer it to a conventional PCR tube or to an AmpliGrid® from Advalytix. The actual lifting of the cells was performed with microglobes that acted as carriers. The microglobes were held with microtweezers and were transferred to the DNA analysis receptacles along with the adhering cells. In a next step, the PCR can be carried out in this receptacle without removing the microglobe. Our method allows a single cell to be isolated directly from evidence material and be made available for forensic DNA analysis.

Empfehlungen der Arbeitsgemeinschaft Forensisch-pädiatrische Diagnostik der Deutschen Gesellschaft für Rechtsmedizin:

ZusammenfassungRegelmäßige weiterführende Untersuchungen im Anschluss an die makroskopische Obduktion von Neugeborenen, Säuglingen oder Kleinkindern sind histologische Untersuchungen sowie toxikologische Analysen mit Blutalkoholbestimmung. DNA-Analysen können im Hinblick auf Täternachweis oder Abstammungsbegutachtung, aber auch als molekulargenetische Untersuchungen auf Erbkrankheiten durchgeführt werden. Außerdem kann der Nachweis einer Stoffwechselstörung aus dem Blut eine Rolle spielen. In seltenen Fällen kann die Forensische Entomologie Relevanz erlangen. Für diese Untersuchungsaufgaben ist die geeignete Asservierung repräsentativer Proben eine zwingende Voraussetzung. Bei Kindern stehen Körperflüssigkeiten oft nur in begrenztem Maße zur Verfügung. Deshalb kann die gleichzeitige Asservierung mehrerer alternativer Proben notwendig sein. Insbesondere bei komplexen Fragestellungen mit unterschiedlichen Untersuchungsanforderungen ist der Einsatz des begrenzten Untersuchungsmaterials unter möglicher Abwägung der Prioritäten als Aufgabe des Rechtsmediziners anzusehen.AbstractFurther regular tests following the macroscopic autopsy of newborns, infants and toddlers are histological examinations as well as toxicological analyses, including a blood alcohol determination. DNA-analyses can be performed with regard to identify a perpetrator or establish genetic relationships, as well as for molecular genetic investigations of hereditary disease. Evidence for a metabolic disorder from blood can also be relevant. In rare cases, forensic entomology can gain importance. The proper storage of representative samples is an essential prerequisite for these examinations. Body fluids are often only available in limited amounts from children. For this reason, a concomitant storage of several alternative samples may be necessary. Particularly in cases with complex questions and diverse testing needs, the use of these limited samples, along with a possible weighing of priorities, must be left to the discretion of the medical examiner.

Investigative strategy for the forensic detection of sperm traces

In a retrospective study, the results from 786 samples of alleged sexual assaults during a 5-year period were evaluated. Of the samples, 758 were from female victims and 28 were from male victims. The material examined during this 5-year period consisted of 561 cotton swabs with swabs taken from the genitals, mouth, anus, or skin surface. In addition, textile products were examined 191 times, paper products 23 times, and other evidentiary materials 11 times. The acid phosphatase (acP) test was performed as a preliminary test for all samples, followed by microscopy after Baecchi staining. DNA analysis was performed on 74 samples following individual court orders. The retrospectively evaluated results from this period indicate that additional tests for the detection of sperm on textiles and paper products are dispensable after a negative acP test. This is different for genital swabs, since sperm could be found microscopically in 3% of cases with a negative acP test, and DNA analysis was also successful. However, an individual investigative strategy has to be determined for each case, as, depending on the structure of the case, the evidence of male DNA on a female victim, or on her clothes, for instance, can also have evidentiary value without microscopic proof for sperm.

Einzelzellen für die forensische DNA-Analyse direkt vom Spurenträger

ZusammenfassungMithilfe eines Mikroskops mit einem digitalen Mikromanipulator werden einzelne Epithelzellen von Spurenträgern gesichert. Die forensische DNA-Analyse der Einzelzellen erfolgt mittels Sequenzierung der hypervariablen Region der mtDNA.AbstractWith a technique using a microscope with a digital micromanipulator, single epithelial cells can be lifted off the surface of material evidence and can be directly made available for forensic DNA-testing. DNA-testing is then accomplished by sequencing the hypervariable region of the mtDNA.