| Zugriffsnummer | 45892 |
| Dokumenttyp | Zeitschriftenartikel |
| Peer Review | mit Peer Review |
| Sprache | Englisch |
| Titel | An assessment of the reproducibility of reverse transcription digital PCR quantification of HIV-1 |
| Autor(in); Institution |
Falak, Samreen; 8.3, Biomedizinische Optik, PTB-Berlin
Busby, Eloise J.; National Measurement Laboratory, LGC, Teddington, UK
O’Sullivan, Denise M.; National Measurement Laboratory, LGC, Teddington, UK
Milavec, Mojca; Department of Biotechnology and Systems Biology, National Institute of Biology, Ljubljana, SLOVENIA
Plauth, Annabell; 8.3, Biomedizinische Optik, PTB-Berlin
Kammel, Martin; INSTAND, Gesellschaft zur Foerderung der Qualitaetssicherung in medizinischen Laboratorien e.V., Düsseldorf, GERMANY
Zeichhardt, Heinz; INSTAND, Gesellschaft zur Foerderung der Qualitaetssicherung in medizinischen Laboratorien e.V., Düsseldorf, GERMANY
Grunert, Hans-Peter; GBD Gesellschaft fuer Biotechnologische Diagnostik mbH, Berlin, GERMANY
Huggett, Jim F.; National Measurement Laboratory, LGC, Teddington, UK
|
| Quelle/Jahr | Methods: 201 (2022), 34 - 40 |
| ISSN | 1046-2023 (PRINT) ; 1095-9130 (ONLINE) |
| DOI | |
| Verlag | Amsterdam: Elsevier |
| Freie Schlagworte | Human immunodeficiency virus type 1 ; Ribonucleic acid ; Digital PCR ; Reverse transcriptase digital PCR ; External quality assessment |
| Zusammenfassung | Viral load monitoring in human immunodeficiency virus type 1 (HIV-1) infection is often performed using reverse transcription quantitative PCR (RT-qPCR) to observe response to treatment and identify the development of resistance. Traceability is achieved using a calibration hierarchy traceable to the International Unit (IU). IU values are determined using consensus agreement derived from estimations by different laboratories. Such a consensus approach is necessary due to the fact that there are currently no reference measurement procedures available that can independently assign a reference value to viral reference materials for molecular in vitro diagnostic tests. Digital PCR (dPCR) is a technique that has the potential to be used for this purpose. In this paper, we investigate the ability of reverse transcriptase dPCR (RT-dPCR) to quantify HIV-1 genomic RNA without calibration. Criteria investigated included the performance of HIV-1 RNA extraction steps, choice of reverse transcription approach and selection of target gene with assays performed in both single and duplex format. We developed a protocol which was subsequently applied by two independent laboratories as part of an external quality assurance (EQA) scheme for HIV-1 genome detection. Our findings suggest that RT- dPCR could be used as reference measurement procedure to aid the value assignment of HIV-1 reference materials to support routine calibration of HIV-1 viral load testing by RT-qPCR. |
| Kostenfreier Zugang | Open Access Hybrid |
| Rechteinformation | CC BY 4.0 ; Creative Commons Attribution 4.0 License |
| Themenbereich der Metrologie | Metrologie in der Medizin |
Zitierung
Falak, S., Macdonald, R., Busby, E. J., O’Sullivan, D. M., Milavec, M., Plauth, A., Kammel, M., Zeichhardt, H., Grunert, H.-P., Kummrow, A., & Huggett, J. F. (2022). An assessment of the reproducibility of reverse transcription digital PCR quantification of HIV-1. Methods, 201, 34–40. https://doi.org/10.1016/j.ymeth.2021.03.006