| Autor: |
Carolin Tumpach, Catherine R. Cochrane, Youry Kim, Jesslyn Ong, Ajantha Rhodes, Thomas A. Angelovich, Melissa J. Churchill, Sharon R. Lewin, Sushama Telwatte, Michael Roche |
| Jazyk: |
angličtina |
| Rok vydání: |
2023 |
| Předmět: |
|
| Zdroj: |
Journal of Virus Eradication, Vol 9, Iss 2, Pp 100335- (2023) |
| Druh dokumentu: |
article |
| ISSN: |
2055-6640 |
| DOI: |
10.1016/j.jve.2023.100335 |
| Popis: |
Quantification of intact proviruses is a critical measurement in HIV cure studies both in vitro and in vivo. The widely adopted ‘intact proviral DNA assay’ (IPDA), designed to discriminate and quantify genetically intact HIV proviruses based on detection of two HIV sequence-specific targets, was originally validated using Bio-Rad's droplet digital PCR technology (ddPCR). Despite its advantages, ddPCR is limited in multiplexing capability (two-channel) and is both labor- and time intensive. To overcome some of these limitations, we utilized a nanowell-based digital PCR platform (dPCR, QIAcuity from Qiagen) which is a fully automated system that partitions samples into nanowells rather than droplets. In this study we adapted the IPDA assay to the QIAcuity platform and assessed its performance relative to ddPCR. The dPCR could differentiate between intact, 5’ defective and 3’ defective proviruses and was sensitive to single HIV copy input. We found the intra-assay and inter-assay variability was within acceptable ranges (with coefficient of variation at or below 10%). When comparing the performance of the IPDA in ex vivo CD4+ T cells from people with HIV on antiretroviral therapy, there was a strong correlation in the quantification of intact (rs = 0.93; p |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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