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...

Descripción completa

Detalles Bibliográficos
Autores principales: 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.
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