Jump to ContentJump to Main Navigation
Show Summary Details
In This Section

Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter / Tate, Jillian R.

12 Issues per year


IMPACT FACTOR 2016: 3.432

CiteScore 2016: 2.21

SCImago Journal Rank (SJR) 2015: 0.873
Source Normalized Impact per Paper (SNIP) 2015: 0.982

Online
ISSN
1437-4331
See all formats and pricing
In This Section
Volume 50, Issue 2 (Feb 2012)

Issues

Strategies of reducing input sample volume for extracting circulating cell-free nuclear DNA and mitochondrial DNA in plasma

Weijie Chen
  • Laboratory for Gynecological Oncology, Department of Biomedicine, University Women’s Hospital, University of Basel, Basel, Switzerland
/ Fengfeng Cai
  • Laboratory for Gynecological Oncology, Department of Biomedicine, University Women’s Hospital, University of Basel, Basel, Switzerland
/ Bei Zhang
  • Laboratory for Gynecological Oncology, Department of Biomedicine, University Women’s Hospital, University of Basel, Basel, Switzerland
/ Xiao Yan Zhong
  • Laboratory for Gynecological Oncology, Department of Biomedicine, University Women’s Hospital, University of Basel, Basel, Switzerland
  • Email:
Published Online: 2011-10-31 | DOI: https://doi.org/10.1515/cclm.2011.773

Abstract

Background: Circulating cell-free (ccf) DNA in blood has been suggested as a potential biomarker in many conditions regarding early diagnosis and prognosis. However, misdiagnosis can result due to the limited DNA resources in Biobank’s plasma samples or insufficient DNA targets from a predominant DNA background in genetic tests. This study explored several strategies for an efficient DNA extraction to increase DNA amount from limited plasma input.

Methods: Ccf plasma DNA was extracted with three different methods, a phenol-chloroform-isoamylalcohol (PCI) method, a High Pure PCR Template Preparation Kit method and a method used for single cell PCR in this group. Subsequently, the total DNA was measured by Nanodrop and the genome equivalents (GE) of the GAPDH housekeeping gene and MTATP 8 gene were measured using a multiplex real-time quantitative PCR for the quantitative assessment of nDNA and mtDNA.

Results: Instead of 400–800 μL (routine input in the laboratory), 50 μLof plasma input enabled the extraction of ccf DNA sufficient for quantitative analysis. Using the PCI method and the kit method, both nDNA and mtDNA could be successfully detected in plasma samples, but nDNA extracted using protocol for single cell PCR was not detectable in 25% of plasma samples. In comparison to the other two methods, the PCI method showed lower DNA purity, but higher concentrations and more GE of nDNA and mtDNA.

Conclusions: The PCI method was more efficient than the other two methods in the extraction of ccf DNA in plasma. Limited plasma is available for ccf DNA analysis.

This article offers supplementary material which is provided at the end of the article.

Keywords: circulating cell-free DNA; mitochondrial DNA; nuclear DNA

About the article

Corresponding author: Xiao Yan Zhong, Laboratory for Gynecological Oncology, Department of Biomedicine, University Women’s Hospital, University of Basel, Hebelstr, 20, 4031 Basel, Switzerland Phone: +41 612 659 248, Fax: +41 612 659 399


Received: 2011-07-07

Accepted: 2011-10-03

Published Online: 2011-10-31

Published in Print: 2012-02-01



Citation Information: Clinical Chemistry and Laboratory Medicine (CCLM), ISSN (Online) 1437-4331, ISSN (Print) 1434-6621, DOI: https://doi.org/10.1515/cclm.2011.773. Export Citation

Supplementary Article Materials

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

[1]
Bernhard Ralla, Carsten Stephan, Sebastian Meller, Dimo Dietrich, Glen Kristiansen, and Klaus Jung
Critical Reviews in Clinical Laboratory Sciences, 2014, Volume 51, Number 4, Page 200

Comments (0)

Please log in or register to comment.
Log in