Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Eberhard, A.
Assessment of pre-analytical factors influencing urinary cell free DNA .
[ Diplomarbeit/Master Thesis (UNI) ] Universität Graz; 2020.
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- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Heitzer Ellen
- Moser Tina
- Altmetrics:
- Abstract:
- Liquid Biopsy refers to the analysis of cell-free DNA (cfDNA) and circulating tumor cells (CTCs) from blood or other body fluids. It is a promising method for the minimally invasive determination and monitoring of the genetic status of tumor patients. The analysis of tumor-derived cell-free circulating tumor DNA (ctDNA) enables a non-invasive analysis of changes in the tumor genome over the entire course of the disease. So far, the main focus of Liquid Biopsy has been on blood. However, cfDNA is also found in other body fluids such as saliva, urine or cerebrospinal fluid (liquor). The use of body fluids other than blood is particularly interesting because there is evidence that taking samples in close proximity to the tumor might increase diagnostic sensitivity. Moreover, in the case of urine, it is a truly non-invasive source of cfDNA. Compared to cfDNA in blood, for which there are already countless studies regarding clinical application, as well as standards and recommendations for pre-analytics, little is known about cfDNA in urine (ucfDNA). In addition, factors such as the high activity of enzymes and possible bacterial contamination make the analysis may complicate the analysis. Although it is known that ucfDNA differs in biology and size from cfDNA in blood, there are no comprehensive data from healthy individuals. Stabilization of blood cells during sample collection has already become a pre-analytical standard. However, there are no recommendations for stabilization of ucfDNA. Therefore, the aim of this study was to investigate the normal state of cfDNA in urine in healthy individuals and to identify any pre-analytical confounding factors and to develop a stable workflow. To this end, urine from healthy controls with and without a stabilizer provided by PreAnalytiX, the PAXgene Blood ccfDNA stabilization reagent, was incubated at RT over a period of 5 days. After ucfDNA extraction, it was analyzed for quantity and integrity. Large fluctuations could be detected between and within volunteers, with women generally showing higher ucfDNA levels than men. Although the available data indicate stabilization of ucfDNA by the PAX reagent, the workflow has to be further optimized to achieve optimal and valid results.