Cargando…
Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA
Substantial efforts to eliminate or reduce latent HIV-1 reservoirs are underway in clinical trials and have created a critical demand for sensitive, accurate, and reproducible tools to evaluate the efficacy of these strategies. Alternative reservoir quantification assays have been developed to circu...
Autores principales: | , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7552071/ https://www.ncbi.nlm.nih.gov/pubmed/32887284 http://dx.doi.org/10.3390/v12090973 |
_version_ | 1783593321618210816 |
---|---|
author | Lungu, Cynthia Procopio, Francesco A. Overmars, Ronald J. Beerkens, Rob J. J. Voermans, Jolanda J. C. Rao, Shringar Prins, Henrieke A. B. Rokx, Casper Pantaleo, Giuseppe van de Vijver, David A. M. C. Mahmoudi, Tokameh Boucher, Charles A. B. Gruters, Rob A. van Kampen, Jeroen J. A. |
author_facet | Lungu, Cynthia Procopio, Francesco A. Overmars, Ronald J. Beerkens, Rob J. J. Voermans, Jolanda J. C. Rao, Shringar Prins, Henrieke A. B. Rokx, Casper Pantaleo, Giuseppe van de Vijver, David A. M. C. Mahmoudi, Tokameh Boucher, Charles A. B. Gruters, Rob A. van Kampen, Jeroen J. A. |
author_sort | Lungu, Cynthia |
collection | PubMed |
description | Substantial efforts to eliminate or reduce latent HIV-1 reservoirs are underway in clinical trials and have created a critical demand for sensitive, accurate, and reproducible tools to evaluate the efficacy of these strategies. Alternative reservoir quantification assays have been developed to circumvent limitations of the quantitative viral outgrowth assay. One such assay is tat/rev induced limiting dilution assay (TILDA), which measures the frequency of CD4+ T cells harboring inducible latent HIV-1 provirus. We modified pre-amplification reagents and conditions (TILDA v2.0) to improve assay execution and first internally validated assay performance using CD4+ T cells obtained from cART-suppressed HIV-1-infected individuals. Detection of tat/rev multiply spliced RNA was not altered by modifying pre-amplification conditions, confirming the robustness of the assay, and supporting the technique’s amenability to limited modifications to ensure better implementation for routine use in clinical studies of latent HIV-1 reservoirs. Furthermore, we cross-validated results of TILDA v2.0 and the original assay performed in two separate laboratories using samples from 15 HIV-1-infected individuals. TILDA and TILDA v2.0 showed a strong correlation (Lin’s Concordance Correlation Coefficient = 0.86). The low inter-laboratory variability between TILDAs performed at different institutes further supports use of TILDA for reservoir quantitation in multi-center interventional HIV-1 Cure trials. |
format | Online Article Text |
id | pubmed-7552071 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-75520712020-10-14 Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA Lungu, Cynthia Procopio, Francesco A. Overmars, Ronald J. Beerkens, Rob J. J. Voermans, Jolanda J. C. Rao, Shringar Prins, Henrieke A. B. Rokx, Casper Pantaleo, Giuseppe van de Vijver, David A. M. C. Mahmoudi, Tokameh Boucher, Charles A. B. Gruters, Rob A. van Kampen, Jeroen J. A. Viruses Brief Report Substantial efforts to eliminate or reduce latent HIV-1 reservoirs are underway in clinical trials and have created a critical demand for sensitive, accurate, and reproducible tools to evaluate the efficacy of these strategies. Alternative reservoir quantification assays have been developed to circumvent limitations of the quantitative viral outgrowth assay. One such assay is tat/rev induced limiting dilution assay (TILDA), which measures the frequency of CD4+ T cells harboring inducible latent HIV-1 provirus. We modified pre-amplification reagents and conditions (TILDA v2.0) to improve assay execution and first internally validated assay performance using CD4+ T cells obtained from cART-suppressed HIV-1-infected individuals. Detection of tat/rev multiply spliced RNA was not altered by modifying pre-amplification conditions, confirming the robustness of the assay, and supporting the technique’s amenability to limited modifications to ensure better implementation for routine use in clinical studies of latent HIV-1 reservoirs. Furthermore, we cross-validated results of TILDA v2.0 and the original assay performed in two separate laboratories using samples from 15 HIV-1-infected individuals. TILDA and TILDA v2.0 showed a strong correlation (Lin’s Concordance Correlation Coefficient = 0.86). The low inter-laboratory variability between TILDAs performed at different institutes further supports use of TILDA for reservoir quantitation in multi-center interventional HIV-1 Cure trials. MDPI 2020-09-02 /pmc/articles/PMC7552071/ /pubmed/32887284 http://dx.doi.org/10.3390/v12090973 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Brief Report Lungu, Cynthia Procopio, Francesco A. Overmars, Ronald J. Beerkens, Rob J. J. Voermans, Jolanda J. C. Rao, Shringar Prins, Henrieke A. B. Rokx, Casper Pantaleo, Giuseppe van de Vijver, David A. M. C. Mahmoudi, Tokameh Boucher, Charles A. B. Gruters, Rob A. van Kampen, Jeroen J. A. Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title | Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title_full | Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title_fullStr | Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title_full_unstemmed | Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title_short | Inter-Laboratory Reproducibility of Inducible HIV-1 Reservoir Quantification by TILDA |
title_sort | inter-laboratory reproducibility of inducible hiv-1 reservoir quantification by tilda |
topic | Brief Report |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7552071/ https://www.ncbi.nlm.nih.gov/pubmed/32887284 http://dx.doi.org/10.3390/v12090973 |
work_keys_str_mv | AT lungucynthia interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT procopiofrancescoa interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT overmarsronaldj interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT beerkensrobjj interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT voermansjolandajc interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT raoshringar interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT prinshenriekeab interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT rokxcasper interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT pantaleogiuseppe interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT vandevijverdavidamc interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT mahmouditokameh interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT bouchercharlesab interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT grutersroba interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda AT vankampenjeroenja interlaboratoryreproducibilityofinduciblehiv1reservoirquantificationbytilda |