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Clinical Chemistry 53: 1349-1357, 2007; 10.1373/clinchem.2006.084004
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(Clinical Chemistry. 2007;53:1349-1357.)
© 2007 American Association for Clinical Chemistry, Inc.

Laboratory Management

EQUAL-qual: A European Program for External Quality Assessment of Genomic DNA Extraction and PCR Amplification

Claudio Orlando1, Paolo Verderio2, Ronald Maatman3, Jan Danneberg3, Simon Ramsden4, Michael Neumaier5, Domenica Taruscio6, Vincenzo Falbo6, Robert Jansen7, Claudia Casini-Raggi1, Francesca Malentacchi1, Ettore Marubini8, Sara Pizzamiglio2, Kris Vernelen9, Jean-Claude Libeer9, Vladimir Palicka10 and Mario Pazzagli1,a

1 Department of Clinical Physiopathology, University of Florence, Florence, Italy.
2 Operative Unit of Medical Statistics and Biometry, Instituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy.
3 Department of Clinical Chemistry, ZiekenhuisGroep Twente, Zilvermeeuw, Almelo, The Netherlands.
4 National Genetics Reference Laboratory, Saint Mary’s Hospital, Manchester, United Kingdom.
5 Institute for Clinical Chemistry, University Hospital Mannheim of the University of Heidelberg, Mannheim, Germany.
6 National Center Rare Diseases, Department of Cell Biology and Neuroscience, Istituto Superiore di Sanità, Rome, Italy.
7 Department of Clinical Chemistry, St. Anna Hospital, Geldrop, The Netherlands.
8 Istituto di Statistica Medica e Biometria, Universitàdegli Studi di Milano, Milan, Italy.
9 Department of Clinical Biology, Institute of Public Health, Brussels, Belgium.
10 Institute for Clinical Chemistry and Diagnostics, School of Medicine, University Hospital Sokolska, Hradec Kralove, Czech Republic.

aAddress correspondence to this author at: Department of Clinical Physiopathology, University of Florence, Florence, Italy. Fax 39-055-4271; e-mail m.pazzagli{at}dfc.unifi.it.

Background: Despite the rapid transition into routine clinical practice of molecular techniques based on PCR, external quality assessment (EQA) is still not widely available. The European Union and European Communities Confederation of Clinical Chemistry have supported the EQUAL project as a series of 3 different EQA programs for the assessment of molecular methods independently from analytes. We present the results from the EQUAL-qual program designed to evaluate the analytical aspects of DNA analysis by means of a conventional qualitative PCR experiment.

Methods: The EQUAL-qual program provided DNA, blood samples, and primer sets to participant laboratories to assess DNA extraction and PCR amplification. We have developed statistical procedures to identify laboratories performing poorly in DNA extraction (quality and quantity), PCR efficiency, and data interpretation after electrophoresis.

Results: An application to participate was obtained from 213 laboratories (from 25 countries), and 175 (82%) of laboratories submitted results for assessment. Questionable results in terms of quality and/or quantity of DNA derived from blood extractions were returned by 27% of laboratories (46 of 166). PCR efficiency showed high variability, with 3% of laboratories (5 of 163) showing a consistently low rate of amplification and 10% (18 of 175) not reporting the expected number of bands of the amplified targets.

Conclusions: The results showed considerable variability in all phases of the experiment. The approach confirms the validity of EQA as a method for evaluating analytical aspects of PCR-based tests.






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Copyright © 2007 by the American Association for Clinical Chemistry.