Performance evaluation of the automated NucliSens easyMAG nucleic acid extraction platform in comparison with QIAamp Mini kit from clinical specimens

Perandin, Francesca; Pollara, P.; Gargiulo, Franco; Bonfanti, Carlo; Manca, N.

doi:10.1016/j.diagmicrobio.2009.02.013

Cited by 20 publications

(18 citation statements)

References 17 publications

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“…samples were randomly allocated to undergo manual or automated extraction, and the analyses were performed on the same days by the same operator who routinely used both methods. Our results are supported by earlier reports which showed that manual extraction methods were comparable [9][10][11] or less sensitive [12][13][14][15] than automated methods for other types of samples and micro-organisms. Besides, automated methods are associated with additional benefits, e.g.…”

Section: Discussionsupporting

confidence: 94%

Optimization of Toxoplasma gondii DNA extraction from amniotic fluid using NucliSENS easyMAG and comparison with QIAamp DNA minikit

Faucher

Miermont

Ranque

et al. 2011

Eur J Clin Microbiol Infect Dis

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Antenatal diagnosis of congenital toxoplasmosis relies on PCR in amniotic fluid. Because parasitic load is often low, DNA extraction must be optimized. Manual methods remain widespread although automated methods appear more effective. This study aimed at optimizing an automated method and at comparing it with a widespread manual method: QIAamp DNA minikit. To optimize NucliSens easyMAG, we evaluated the addition of proteinase K pre-treatment and the increase of the amount of silica particles used for the extraction. The optimized method was then compared to QIAamp DNA minikit on samples containing less than 25 tachyzoites/ml. NucliSens easyMAG DNA yield was improved after proteinase K pre-treatment (p < 0.01), but not with a higher silica particle input. The optimized method yielded more positive PCRs than the manual method, especially for samples containing 5 tachyzoites/ml or less (71% vs 26%, p < 10(-4)). The DNA amount in samples found positive by PCR was higher after optimized automated extraction than after manual extraction (p < 10(-4)). Proteinase K pre-treatment should be added to extract DNA from amniotic fluid using NucliSens easyMAG. Using this optimized automated method rather than manual methods would improve the sensitivity of Toxoplasma PCR and simplify the daily workflow.

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Section: Discussionsupporting

confidence: 94%

Optimization of Toxoplasma gondii DNA extraction from amniotic fluid using NucliSENS easyMAG and comparison with QIAamp DNA minikit

Faucher

Miermont

Ranque

et al. 2011

Eur J Clin Microbiol Infect Dis

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“…However, as dPCR platforms are considered to be relatively robust to potential inhibitory substances that might have remained during the DNA extraction [11, 12], it is likely that the differences in the estimated mean DNA copy number concentration between the WVM units were mostly caused by variable DNA recovery upon extraction. This is in agreement with previously reported data with HCMV, where intermediate variability was noted between extraction replicates analysed by dPCR within a single laboratory [20, 31]. With the QX100 system, the differences between the WVM units had higher statistical significance in comparison to the Biomark 37K array.…”

Section: Resultssupporting

confidence: 92%

“…With the low filling-volume related uncertainty and high stability of the WVM units that are claimed by the manufacturer, it is reasonable to assume that the inter-unit variability was introduced during the DNA extraction, as the DNA was locally extracted from each individual WVM unit. This is in agreement with previous studies where up to 50% difference was noted between DNA-extraction replicates quantified using the same qPCR assay [30, 31]. With PCR-based DNA quantification, estimation of DNA copy number concentration can be influenced by variable DNA recovery and/or insufficient removal of PCR inhibitors during DNA extractions [32].…”

Section: Resultssupporting

confidence: 91%

“…In contrast to the centrally prepared gDNA test material, the WVM required local DNA extraction before DNA quantification. DNA extraction has already been demonstrated to introduce an additional variability (CV up to 50%) due to differences in DNA recoveries from extraction columns of the same manual extraction kit used by one operator within one laboratory [20, 30, 31]. To the best of our knowledge, no inter-laboratory assessment of the same DNA extraction method for quantification of viral DNA has been performed.…”

Section: Resultsmentioning

confidence: 99%

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Inter-laboratory assessment of different digital PCR platforms for quantification of human cytomegalovirus DNA

Pavšič

Devonshire²,

Blejec

et al. 2017

Anal Bioanal Chem

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Quantitative PCR (qPCR) is an important tool in pathogen detection. However, the use of different qPCR components, calibration materials and DNA extraction methods reduces comparability between laboratories, which can result in false diagnosis and discrepancies in patient care. The wider establishment of a metrological framework for nucleic acid tests could improve the degree of standardisation of pathogen detection and the quantification methods applied in the clinical context. To achieve this, accurate methods need to be developed and implemented as reference measurement procedures, and to facilitate characterisation of suitable certified reference materials. Digital PCR (dPCR) has already been used for pathogen quantification by analysing nucleic acids. Although dPCR has the potential to provide robust and accurate quantification of nucleic acids, further assessment of its actual performance characteristics is needed before it can be implemented in a metrological framework, and to allow adequate estimation of measurement uncertainties. Here, four laboratories demonstrated reproducibility (expanded measurement uncertainties below 15%) of dPCR for quantification of DNA from human cytomegalovirus, with no calibration to a common reference material. Using whole-virus material and extracted DNA, an intermediate precision (coefficients of variation below 25%) between three consecutive experiments was noted. Furthermore, discrepancies in estimated mean DNA copy number concentrations between laboratories were less than twofold, with DNA extraction as the main source of variability. These data demonstrate that dPCR offers a repeatable and reproducible method for quantification of viral DNA, and due to its satisfactory performance should be considered as candidate for reference methods for implementation in a metrological framework.Electronic supplementary materialThe online version of this article (doi:10.1007/s00216-017-0206-0) contains supplementary material, which is available to authorized users.

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“…These observations correspond with recent studies in which the comparison of manual and automated CMV DNA extraction methods yielded conflicting results. 5,7,[24][25][26] Recently, the mean CMV DNA concentration obtained after extraction on the QIAamp DNA Mini Kit (Qiagen), MagNA Pure LC (Roche Diagnostics), or easyMAG platform (bioMerieux) was found to vary up to 2.3-fold. 26 In this study, the mean CMV DNA concentrations varied up to 5.5 fold in quantification between the highest (Qiasymphony) and lowest extraction performance (m2000sp) but only few differences were found to be greater than 1.0 log when comparing results obtained from individual samples.…”

Section: Discussionmentioning

confidence: 99%

Extraction of viral nucleic acids: Comparison of five automated nucleic acid extraction platforms

Verheyen

Kaiser

Bozic

et al. 2012

Journal of Clinical Virology

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Performance evaluation of the automated NucliSens easyMAG nucleic acid extraction platform in comparison with QIAamp Mini kit from clinical specimens

Cited by 20 publications

References 17 publications

Optimization of Toxoplasma gondii DNA extraction from amniotic fluid using NucliSENS easyMAG and comparison with QIAamp DNA minikit

Optimization of Toxoplasma gondii DNA extraction from amniotic fluid using NucliSENS easyMAG and comparison with QIAamp DNA minikit

Inter-laboratory assessment of different digital PCR platforms for quantification of human cytomegalovirus DNA

Extraction of viral nucleic acids: Comparison of five automated nucleic acid extraction platforms

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