Extraction of viral nucleic acids: comparison of five automated nucleic acid extraction platforms
- PMID: 22503856
- DOI: 10.1016/j.jcv.2012.03.008
Extraction of viral nucleic acids: comparison of five automated nucleic acid extraction platforms
Abstract
Background: Nucleic acid extraction has a major impact on the reliability of results in routine molecular diagnostics. Optimal isolation of nucleic acids and removal of inhibitors are essential.
Objectives: This study compares five different automated extraction platforms for the extraction of norovirus RNA from stool and cytomegalovirus (CMV) DNA from plasma samples.
Study design: Norovirus positive stool samples and CMV positive plasma samples were aliquoted and analyzed using five different automated platforms (easyMAG, bioMerieux; m2000sp, Abbott; MagNA Pure LC 2.0, Roche; QiaSymphony, Qiagen; sample preparation module of the VERSANT kPCR Molecular System, Siemens). Similar sample input and output volumes, the identical real-time PCR cycler, and the identical assays for amplification and detection for norovirus RNA and CMV DNA, respectively, were chosen.
Results: Of 39 stool samples, 36 tested positive for norovirus RNA with all extraction platforms. The three discrepant samples showed inhibition after extraction with at least one platform. Only with the VERSANT platform all samples tested positive for both the target RNA and the internal controls. Of 42 plasma samples, 27 gave quantifiable results for CMV DNA with all extraction platforms. There was significant variance between viral concentrations when different extraction platforms were compared. The majority of the 15 discrepant samples showed low viral concentrations. The internal control of the CMV assay gave positive results for all samples tested below the limit of quantification.
Conclusions: The five automated extraction platforms yielded comparable results. However, the extraction performance was found to be impaired by inhibitory substances in stool samples.
Copyright © 2012 Elsevier B.V. All rights reserved.
Similar articles
-
Comparative evaluation of the QIAGEN QIAsymphony® SP system and bioMérieux NucliSens easyMAG automated extraction platforms in a clinical virology laboratory.J Clin Virol. 2011 Dec;52(4):339-43. doi: 10.1016/j.jcv.2011.08.016. Epub 2011 Oct 20. J Clin Virol. 2011. PMID: 22014618
-
Performance evaluation of the automated NucliSens easyMAG nucleic acid extraction platform in comparison with QIAamp Mini kit from clinical specimens.Diagn Microbiol Infect Dis. 2009 Jun;64(2):158-65. doi: 10.1016/j.diagmicrobio.2009.02.013. Diagn Microbiol Infect Dis. 2009. PMID: 19500527
-
Improved quantitative PCR protocols for adenovirus and CMV with an internal inhibition control system and automated nucleic acid isolation.J Med Virol. 2012 Jun;84(6):890-6. doi: 10.1002/jmv.23285. J Med Virol. 2012. PMID: 22499011
-
Quantitative TaqMan assay for the detection and monitoring of cytomegalovirus infection in organ transplant patients.Methods Mol Biol. 2006;335:147-56. doi: 10.1385/1-59745-069-3:147. Methods Mol Biol. 2006. PMID: 16785626 Review.
-
Quantitative molecular methods in virology.Arch Virol. 1995;140(9):1523-39. doi: 10.1007/BF01322527. Arch Virol. 1995. PMID: 7487485 Review.
Cited by
-
Performance and Application of 16S rRNA Gene Cycle Sequencing for Routine Identification of Bacteria in the Clinical Microbiology Laboratory.Clin Microbiol Rev. 2020 Sep 9;33(4):e00053-19. doi: 10.1128/CMR.00053-19. Print 2020 Sep 16. Clin Microbiol Rev. 2020. PMID: 32907806 Free PMC article. Review.
-
Evaluation of DNA Extraction Methods for Reliable Quantification of Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa.Biosensors (Basel). 2023 Apr 6;13(4):463. doi: 10.3390/bios13040463. Biosensors (Basel). 2023. PMID: 37185538 Free PMC article.
-
Increased frequency of JC-polyomavirus detection in rheumatoid arthritis patients treated with multiple biologics.Med Microbiol Immunol. 2015 Oct;204(5):613-8. doi: 10.1007/s00430-015-0390-5. Epub 2015 Feb 13. Med Microbiol Immunol. 2015. PMID: 25678083
-
Comparison of Luminex xTAG® RVP fast assay and real time RT-PCR for the detection of respiratory viruses in adults with community-acquired pneumonia.J Med Virol. 2016 Jul;88(7):1173-9. doi: 10.1002/jmv.24463. Epub 2016 Feb 2. J Med Virol. 2016. PMID: 27061405 Free PMC article.
-
Does Size Matter? Comparison of Extraction Yields for Different-Sized DNA Fragments by Seven Different Routine and Four New Circulating Cell-Free Extraction Methods.J Clin Microbiol. 2018 Nov 27;56(12):e01061-18. doi: 10.1128/JCM.01061-18. Print 2018 Dec. J Clin Microbiol. 2018. PMID: 30282788 Free PMC article.
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Medical
