Evaluation of Preanalytical Conditions and Implementation of Quality Control Steps for Reliable Gene Expression and DNA Methylation Analyses in Liquid Biopsies

Clinical Chemistry
Martha ZavridouEvi S Lianidou

Abstract

Liquid biopsy provides important information for the prognosis and treatment of cancer patients. In this study, we evaluated the effects of preanalytical conditions on gene expression and DNA methylation analyses in liquid biopsies. We tested the stability of circulating tumor cell (CTC) messenger RNA by spiking MCF-7 cells in healthy donor peripheral blood (PB) drawn into 6 collection-tube types with various storage conditions. CTCs were enriched based on epithelial cell adhesion molecule positivity, and RNA was isolated followed by cDNA synthesis. Gene expression was quantified using RT-quantitative PCR for CK19 and B2M. We evaluated the stability of DNA methylation in plasma under different storage conditions by spiking DNA isolated from MCF-7 cells in healthy donor plasma. Two commercially available sodium bisulfite (SB)-conversion kits were compared, in combination with whole genome amplification (WGA), to evaluate the stability of SB-converted DNA. SB-converted DNA samples were analyzed by real-time methylation-specific PCR (MSP) for ACTB, SOX17, and BRMS1. Quality control was assessed using Levey-Jennings graphs. RNA-based analysis in CTCs is severely impeded by the preservatives used in many PB collection tubes (except ...Continue Reading

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Citations

May 13, 2020·Critical Reviews in Clinical Laboratory Sciences·Vida UngererStefan Holdenrieder
Nov 2, 2019·Critical Reviews in Clinical Laboratory Sciences·Eleni Tzanikou, Evi Lianidou
Jul 18, 2020·Current Opinion in Oncology·Lisanne F van DesselMartijn P Lolkema
Jul 21, 2020·Journal of Analytical Toxicology·Kelly Francisco da CunhaJose Luiz Costa
Oct 20, 2020·Epigenetics : Official Journal of the DNA Methylation Society·Ruben Van PaemelKatleen De Preter
Jan 26, 2021·Biopreservation and Biobanking·Smadar Kahana-EdwinJonathan Karpelowsky
Nov 13, 2019·Molecular Aspects of Medicine·Edward Geeurickx, An Hendrix
Feb 23, 2021·Frontiers in Cell and Developmental Biology·Aitor Rodriguez-CasanovaAngel Diaz-Lagares
Apr 28, 2021·Journal of Experimental & Clinical Cancer Research : CR·Yan LiChuanyong Guo
Aug 15, 2021·Journal of Clinical Laboratory Analysis·Xuan-Yu HuZhi-Qiang Ling

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