Harmonisation of multi-centre real-time reverse-transcribed PCR results of a candidate prognostic marker in breast cancer: an EU-FP6 supported study of members of the EORTC - PathoBiology Group.

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Publication year
2009Source
European Journal of Cancer, 45, 1, (2009), pp. 74-81ISSN
Publication type
Article / Letter to editor

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Organization
Chemical Endocrinology
Radiation Oncology
Laboratory of Genetic, Endocrine and Metabolic Diseases
Journal title
European Journal of Cancer
Volume
vol. 45
Issue
iss. 1
Page start
p. 74
Page end
p. 81
Subject
ONCOL 1: Hereditary cancer and cancer-related syndromes; ONCOL 3: Translational research; ONCOL 5: Aetiology, screening and detectionAbstract
AIM: Assessment of intra- and inter-laboratory variation in multi-centre real-time reverse-transcribed PCR (qRT-PCR)-based mRNA quantification of a prognostic marker in breast cancer using external quality assurance (EQA). METHODS: A questionnaire on the methodologies used and EQA calibrators were sent to 5 participating laboratories from 4 European countries, which measured mRNA levels of PITX2 splice variants and reference genes by qRT-PCR. RESULTS: Differences in the methodology included PCR quantification methodology and equipment, RNA extraction and cDNA synthesis procedures. The intra-laboratory coefficient of variation (CV) ranged from 5 to 23%, and the inter-laboratory CV ranged from 17 to 30%. The inter-laboratory CV was reduced to 13% by using prediluted calibrators and by harmonising the data in the central QA laboratory. Additional normalisation using reference genes did not decrease the variation further. CONCLUSIONS: Both externally provided calibrators and centralised harmonisation are required to reduce the intra-laboratory variation in multi-centre qRT-PCR results to an acceptable level.
This item appears in the following Collection(s)
- Academic publications [227883]
- Electronic publications [107369]
- Faculty of Medical Sciences [86219]
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