Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Maier, BK.
Extraction of CMV DNA out of EDTA whole blood samples: comparison of automated platforms
[ Diplomarbeit/Master Thesis ] Medical University of Graz; 2011. pp.50.
[OPEN ACCESS]
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- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Kessler Harald
- Altmetrics:
- Abstract:
- Background: Detection and quantification of cytomegalovirus (CMV) DNA by real-time PCR (qPCR) is the standard method to detect early CMV infection. Because the qPCR technology is unlikely to be further improved, the current focus is on optimization of nucleic acid extraction. Objectives: To evaluate and compare automated platforms useful for extraction of CMV DNA from different sample types including EDTA whole blood, plasma, urine, and bronchoalveolar lavage. To compare results obtained from different real-time PCR systems. To compare time-to-result when employing different assays. Materials & Methods: Twenty clinical samples obtained from patients with CMV infection including 5 EDTA whole blood, 5 plasma, 5 urine, and 5 bronchoalveolar lavage samples were analyzed on 4 different automated extraction platforms (easyMAG, QIAsymphony, MagNA Pure LC 2.0, VERSANT kPCR SP). Nucleic acid extracts were amplified and detected on 2 different qPCR instruments, the LightCycler 480 II and the VERSANT kPCR AD. Results: When the VERSANT kPCR SP instrument in conjunction with the VERSANT kPCR AD was compared to the alternative extraction systems, significantly higher CMV DNA concentrations were obtained for ETDA whole blood and plasma samples (p<0.05). When comparing the two amplification systems, the LightCycler 480 II gave significantly higher values for the majority of samples. The VERSANT kPCR SP has the longest total time-to-result. Conclusion: With each assay and sample type, consistent results could be generated within an acceptable timeframe. It should be considered that a statistically significantly higher CMV DNA concentration does not necessarily imply clinical significance.