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Validation guidelines for PCR workflows in bioterrorism preparedness, food safety and forensics

Hedman, Johannes LU ; Lavander, Moa; Salomonsson, Emelie Näslund; Jinnerot, Tomas; Boiso, Lina; Magnusson, Bertil and Rådström, Peter LU (2018) In Accreditation and Quality Assurance p.1-12
Abstract

The polymerase chain reaction (PCR) is the backbone of contemporary DNA/RNA analysis, ideally enabling detection of one or just a few target molecules. However, when analysing food or forensic samples the analytical procedure is often challenged by low amounts of poor quality template molecules and complex matrices. Applying optimised and validated methods in all steps of the analysis workflow, i.e. sampling, sample treatment, DNA/RNA extraction and PCR (including reverse transcription for RNA analysis), is thus necessary to ensure the reliability of analysis. In this paper, we describe how in-house validation can be performed for the different modules of the diagnostic PCR process, providing practical examples as tools for laboratories... (More)

The polymerase chain reaction (PCR) is the backbone of contemporary DNA/RNA analysis, ideally enabling detection of one or just a few target molecules. However, when analysing food or forensic samples the analytical procedure is often challenged by low amounts of poor quality template molecules and complex matrices. Applying optimised and validated methods in all steps of the analysis workflow, i.e. sampling, sample treatment, DNA/RNA extraction and PCR (including reverse transcription for RNA analysis), is thus necessary to ensure the reliability of analysis. In this paper, we describe how in-house validation can be performed for the different modules of the diagnostic PCR process, providing practical examples as tools for laboratories in their planning of validation studies. The focus is analysis of heterogeneous samples with interfering matrices, with relevance in food testing, forensic DNA analysis, bioterrorism preparedness and veterinary medicine. Our objective is to enable rational in-house validation for reliable and swift quality assurance when results are urgent, for example in the event of a crisis such as a foodborne outbreak or a crime requiring the analysis of a large number of diverse samples. To that end, we explain the performance characteristics associated with method validation from a PCR and biological sample matrix perspective and suggest which characteristics to investigate depending on the type of method to be validated. Also, we include a modular approach to validation within the PCR workflow, aiming at efficient validation and a flexible use of methods.

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author
organization
publishing date
type
Contribution to journal
publication status
epub
subject
keywords
Digital PCR, Forensic DNA analysis, ISO/IEC 17025, Pre-PCR processing, qPCR, Quality assurance, Validation
in
Accreditation and Quality Assurance
pages
12 pages
publisher
Springer
external identifiers
  • scopus:85048025575
ISSN
0949-1775
DOI
language
English
LU publication?
yes
id
f0963f87-e08c-4afa-89e0-d3c092213862
date added to LUP
2018-06-11 11:36:29
date last changed
2018-06-11 11:36:29
@article{f0963f87-e08c-4afa-89e0-d3c092213862,
  abstract     = {<p>The polymerase chain reaction (PCR) is the backbone of contemporary DNA/RNA analysis, ideally enabling detection of one or just a few target molecules. However, when analysing food or forensic samples the analytical procedure is often challenged by low amounts of poor quality template molecules and complex matrices. Applying optimised and validated methods in all steps of the analysis workflow, i.e. sampling, sample treatment, DNA/RNA extraction and PCR (including reverse transcription for RNA analysis), is thus necessary to ensure the reliability of analysis. In this paper, we describe how in-house validation can be performed for the different modules of the diagnostic PCR process, providing practical examples as tools for laboratories in their planning of validation studies. The focus is analysis of heterogeneous samples with interfering matrices, with relevance in food testing, forensic DNA analysis, bioterrorism preparedness and veterinary medicine. Our objective is to enable rational in-house validation for reliable and swift quality assurance when results are urgent, for example in the event of a crisis such as a foodborne outbreak or a crime requiring the analysis of a large number of diverse samples. To that end, we explain the performance characteristics associated with method validation from a PCR and biological sample matrix perspective and suggest which characteristics to investigate depending on the type of method to be validated. Also, we include a modular approach to validation within the PCR workflow, aiming at efficient validation and a flexible use of methods.</p>},
  author       = {Hedman, Johannes and Lavander, Moa and Salomonsson, Emelie Näslund and Jinnerot, Tomas and Boiso, Lina and Magnusson, Bertil and Rådström, Peter},
  issn         = {0949-1775},
  keyword      = {Digital PCR,Forensic DNA analysis,ISO/IEC 17025,Pre-PCR processing,qPCR,Quality assurance,Validation},
  language     = {eng},
  month        = {06},
  pages        = {1--12},
  publisher    = {Springer},
  series       = {Accreditation and Quality Assurance},
  title        = {Validation guidelines for PCR workflows in bioterrorism preparedness, food safety and forensics},
  url          = {http://dx.doi.org/},
  year         = {2018},
}