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Establishing PNB-qPCR for quantifying minimal ctDNA concentrations during tumour resection
1Department of Sports Medicine, Rehabilitation and Disease Prevention, Faculty of Social Science, Media and Sport, Johannes Gutenberg-University Mainz, Mainz, Germany.
Scientific Reports
|August 23, 2017
Summary
This study introduces PNB-qPCR to detect circulating tumor DNA (ctDNA) in plasma. The method reveals ctDNA and cell-free DNA (cfDNA) release patterns after tumor surgery.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Liquid biopsies offer a non-invasive alternative to tissue biopsies for cancer research.
- Detecting circulating tumor DNA (ctDNA) in plasma aids in monitoring tumor status, prognosis, and therapy response.
Purpose of the Study:
- To develop a highly sensitive method for detecting and quantifying minute amounts of ctDNA in limited plasma samples.
- To characterize the kinetics of ctDNA and total cell-free DNA (cfDNA) following tumor resection.
Main Methods:
- Established PNB-qPCR (Pooled, Nested, WT-Blocking qPCR), a nested qPCR technique.
- Combined ARMS primers, blocking primers, LNA probes, and pooling for sensitive quantification of KRAS mutations.
- Analyzed ctDNA and cfDNA levels in plasma during and after tumor resection.
Main Results:
- PNB-qPCR accurately detected and quantified ctDNA from small plasma volumes.
- Total cfDNA increased during the surgical and regenerative process.
- ctDNA levels specifically rose immediately after tumor resection and around three days post-surgery.
Conclusions:
- PNB-qPCR is a sensitive method for analyzing ctDNA in plasma.
- Surgery impacts ctDNA and cfDNA release differently.
- This study provides novel insights into ctDNA dynamics after tumor resection.
