Detection of Cell-Free DNA in Blood Plasma Samples of Cancer Patients
| Autor: | Milana Frenkel-Morgenstern, Dorith Raviv Shay, Vikrant Palande |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
General Chemical Engineering
Buffy coat General Biochemistry Genetics and Molecular Biology chemistry.chemical_compound Neoplasms Biopsy medicine Biomarkers Tumor Humans Fluorometry Liquid biopsy General Immunology and Microbiology medicine.diagnostic_test business.industry Genome Human General Neuroscience Liquid Biopsy Cancer DNA Neoplasm Glioma medicine.disease genomic DNA Cell-free fetal DNA chemistry Cancer cell Cancer research business Cell-Free Nucleic Acids DNA |
| Zdroj: | Journal of visualized experiments : JoVE. (163) |
| ISSN: | 1940-087X |
| Popis: | Identifying mutations in tumors of cancer patients is a very important step in disease management. These mutations serve as biomarkers for tumor diagnosis as well as for the treatment selection and its response in cancer patients. The current gold standard method for detecting tumor mutations involves a genetic test of tumor DNA by means of tumor biopsies. However, this invasive method is difficult to be performed repeatedly as a follow-up test of the tumor mutational repertoire. Liquid biopsy is a new and emerging technique for detecting tumor mutations as an easy-to-use and non-invasive biopsy approach. Cancer cells multiply rapidly. In parallel, numerous cancer cells undergo apoptosis. Debris from these cells are released into a patient's circulatory system, together with finely fragmented DNA pieces, called cell-free DNA (cfDNA) fragments, which carry tumor DNA mutations. Therefore, for identifying cfDNA based biomarkers using liquid biopsy technique, blood samples are collected from the cancer patients, followed by the separation of plasma and buffy coat. Next, plasma is processed for the isolation of cfDNA, and the respective buffy coat is processed for the isolation of a patient's genomic DNA. Both nucleic acid samples are then checked for their quantity and quality; and analyzed for mutations using next-generation sequencing (NGS) techniques. In this manuscript, we present a detailed protocol for liquid biopsy, including blood collection, plasma, and buffy coat separation, cfDNA and germline DNA extraction, quantification of cfDNA or germline DNA, and cfDNA fragment enrichment analysis. |
| Databáze: | OpenAIRE |
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