Dataset Information

PowerPlex® fusion 6C system: internal validation study.

ABSTRACT: This work is aimed at describing the proceedings and parameters used to validate PowerPlex® Fusion 6C System, the polymerase chain reaction (PCR) amplification kit by Promega, for posterior implementation in the laboratorial routine of the Forensic Genetic Service. The PowerPlex® Fusion 6C System allows multiplex PCR, through simultaneous amplification and posterior detection by fluorescence of 27 loci. Characterization of the kit was made according to the laboratory's internal validation procedure based on validation guidelines from Scientific Working Group on DNA Analysis Methods. Some parameters were evaluated, such as specificity, analytical thresholds, sensitivity, precision, mixture studies, DNA control samples, a proficiency test and changes in the PCR-based procedures: final reaction volume and cycle number, changes in the reaction mixture for direct amplification. This kit proved to be very robust and the results are in concordance with previous developmental validation by the manufacturer. In some parameters, the results were better than expected.

SUBMITTER: Boavida A

PROVIDER: S-EPMC6197087 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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PowerPlex® fusion 6C system: internal validation study.

Boavida Ana A Bogas Vanessa V Sampaio Lisa L Gouveia Nair N Porto Maria J MJ Corte-Real Francisco F

Forensic sciences research 20180209 2

This work is aimed at describing the proceedings and parameters used to validate PowerPlex® Fusion 6C System, the polymerase chain reaction (PCR) amplification kit by Promega, for posterior implementation in the laboratorial routine of the Forensic Genetic Service. The PowerPlex® Fusion 6C System allows multiplex PCR, through simultaneous amplification and posterior detection by fluorescence of 27 loci. Characterization of the kit was made according to the laboratory's internal validation proced ...[more]

PMID: 30483661

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Project description:Combining diagnostic specimens into pools has been considered as a strategy to augment throughput, decrease turnaround time, and leverage resources. This study utilized a multi-parametric approach to assess optimum pool size, impact of automation, and effect of nucleic acid amplification chemistries on the detection of SARS-CoV-2 RNA in pooled samples for surveillance testing on the Hologic Panther Fusion® System. Dorfman pooled testing was conducted with previously tested SARS-CoV-2 nasopharyngeal samples using Hologic's Aptima® and Panther Fusion® SARS-CoV-2 Emergency Use Authorization assays. A manual workflow was used to generate pool sizes of 5:1 (five samples: one positive, four negative) and 10:1. An automated workflow was used to generate pool sizes of 3:1, 4:1, 5:1, 8:1 and 10:1. The impact of pool size, pooling method, and assay chemistry on sensitivity, specificity, and lower limit of detection (LLOD) was evaluated. Both the Hologic Aptima® and Panther Fusion® SARS-CoV-2 assays demonstrated >85% positive percent agreement between neat testing and pool sizes ≤5:1, satisfying FDA recommendation. Discordant results between neat and pooled testing were more frequent for positive samples with CT>35. Fusion® CT (cycle threshold) values for pooled samples increased as expected for pool sizes of 5:1 (CT increase of 1.92-2.41) and 10:1 (CT increase of 3.03-3.29). The Fusion® assay demonstrated lower LLOD than the Aptima® assay for pooled testing (956 vs 1503 cp/mL, pool size of 5:1). Lowering the cut-off threshold of the Aptima® assay from 560 kRLU (manufacturer's setting) to 350 kRLU improved the assay sensitivity to that of the Fusion® assay for pooled testing. Both Hologic's SARS-CoV-2 assays met the FDA recommended guidelines for percent positive agreement (>85%) for pool sizes ≤5:1. Automated pooling increased test throughput and enabled automated sample tracking while requiring less labor. The Fusion® SARS-CoV-2 assay, which demonstrated a lower LLOD, may be more appropriate for surveillance testing.

Project description:Background. Auditory event-related potentials (ERPs) have proved useful in investigating the role of auditory processing in cognitive disorders such as developmental dyslexia, specific language impairment (SLI), attention deficit hyperactivity disorder (ADHD), schizophrenia, and autism. However, laboratory recordings of auditory ERPs can be lengthy, uncomfortable, or threatening for some participants - particularly children. Recently, a commercial gaming electroencephalography (EEG) system has been developed that is portable, inexpensive, and easy to set up. In this study we tested if auditory ERPs measured using a gaming EEG system (Emotiv EPOC(®), www.emotiv.com) were equivalent to those measured by a widely-used, laboratory-based, research EEG system (Neuroscan). Methods. We simultaneously recorded EEGs with the research and gaming EEG systems, whilst presenting 21 adults with 566 standard (1000 Hz) and 100 deviant (1200 Hz) tones under passive (non-attended) and active (attended) conditions. The onset of each tone was marked in the EEGs using a parallel port pulse (Neuroscan) or a stimulus-generated electrical pulse injected into the O1 and O2 channels (Emotiv EPOC(®)). These markers were used to calculate research and gaming EEG system late auditory ERPs (P1, N1, P2, N2, and P3 peaks) and the mismatch negativity (MMN) in active and passive listening conditions for each participant. Results. Analyses were restricted to frontal sites as these are most commonly reported in auditory ERP research. Intra-class correlations (ICCs) indicated that the morphology of the research and gaming EEG system late auditory ERP waveforms were similar across all participants, but that the research and gaming EEG system MMN waveforms were only similar for participants with non-noisy MMN waveforms (N = 11 out of 21). Peak amplitude and latency measures revealed no significant differences between the size or the timing of the auditory P1, N1, P2, N2, P3, and MMN peaks. Conclusions. Our findings suggest that the gaming EEG system may prove a valid alternative to laboratory ERP systems for recording reliable late auditory ERPs (P1, N1, P2, N2, and the P3) over the frontal cortices. In the future, the gaming EEG system may also prove useful for measuring less reliable ERPs, such as the MMN, if the reliability of such ERPs can be boosted to the same level as late auditory ERPs.

Dataset Information

PowerPlex® fusion 6C system: internal validation study.

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PowerPlex® fusion 6C system: internal validation study.

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