Real-time PCR applications in haemostasis and transfusion medicine
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This thesis deals with the development and application of novel real-time PCR assays in the field of haemostasis and transfusion medicine. Chapter 1 gives an general overview of real-time PCR and its current applications in haemotherapy. The development and validation of a real-time PCR assay for the detection of Parvovirus B19 DNA in blood donations is described in chapter 2. This sensitive and internally controlled PCR assay fulfils all criteria for the routine screening of predominantly B19 negative blood donations. A real-time PCR platform for the quantification of tissue factor (TF) mRNA in circulating blood monocytes is presented in chapter 3. To ensure reliable test results, a blood sampling system that includes a RNA stabilizing additive was evaluated. Utilizing the described TF real-time PCR assay we were able to demonstrate that desmopressin acetate (DDAVP) is able to induce monocytic tissue factor gene expression in vivo (chapter 4). By the application of different real-time PCR assays for absolute quantification of chosen marker transcripts, it was possible to validate the so-called SMART procedure for global amplification of platelet mRNA sequences applied for subsequent gene expression analysis (chapter 5). As described in chapter 6, different 5 -nuclease assays were used for the allelic discrimination of thrombophilic risk factors in patients with dural arteriovenous fistuals (DAVFs). In summary, the real-time 5 -nuclease assays described in this thesis showed to be valuable methods that strongly improved the possibilities of molecular analysis in haemostasis and transfusion medicine.Verknüpfung zu Publikationen oder weiteren Datensätzen
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Infusion Therapy and Transfusion Medicine, 29 (2002), S. 254-258 Journal of Neurology, 249 (2002), S. 680-682; Clinical Chemistry, 50 (2004), S. 245-247, S. 2271-2287