Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.

The analysis of the infectious titer of the lentiviral vector samples obtained during upstream and downstream processing is of major importance, however, also the most challenging method to be performed. Currently established methods like flow cytometry or qPCR lack the capability of enabling high t...

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Autores principales:	Jennifer J Labisch, G Philip Wiese, Kalpana Barnes, Franziska Bollmann, Karl Pflanz
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/e96cb8de96504815a35df8cfd66633ec
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spelling	oai:doaj.org-article:e96cb8de96504815a35df8cfd66633ec2021-12-02T20:09:10ZInfectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.1932-620310.1371/journal.pone.0254739https://doaj.org/article/e96cb8de96504815a35df8cfd66633ec2021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0254739https://doaj.org/toc/1932-6203The analysis of the infectious titer of the lentiviral vector samples obtained during upstream and downstream processing is of major importance, however, also the most challenging method to be performed. Currently established methods like flow cytometry or qPCR lack the capability of enabling high throughput sample processing while they require a lot of manual handling. To address this limitation, we developed an immunological real-time imaging method to quantify the infectious titer of anti-CD19 CAR lentiviral vectors with a temporal readout using the Incucyte® S3 live-cell analysis system. The infective titers determined with the Incucyte® approach when compared with the flow cytometry-based assay had a lower standard deviation between replicates and a broader linear range. A major advantage of the method is the ability to obtain titer results in real-time, enabling an optimal readout time. The presented protocol significantly decreased labor and increased throughput. The ability of the assay to process high numbers of lentiviral samples in a high throughput manner was proven by performing a virus stability study, demonstrating the effects of temperature, salt, and shear stress on LV infectivity.Jennifer J LabischG Philip WieseKalpana BarnesFranziska BollmannKarl PflanzPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 7, p e0254739 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jennifer J Labisch G Philip Wiese Kalpana Barnes Franziska Bollmann Karl Pflanz Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
description	The analysis of the infectious titer of the lentiviral vector samples obtained during upstream and downstream processing is of major importance, however, also the most challenging method to be performed. Currently established methods like flow cytometry or qPCR lack the capability of enabling high throughput sample processing while they require a lot of manual handling. To address this limitation, we developed an immunological real-time imaging method to quantify the infectious titer of anti-CD19 CAR lentiviral vectors with a temporal readout using the Incucyte® S3 live-cell analysis system. The infective titers determined with the Incucyte® approach when compared with the flow cytometry-based assay had a lower standard deviation between replicates and a broader linear range. A major advantage of the method is the ability to obtain titer results in real-time, enabling an optimal readout time. The presented protocol significantly decreased labor and increased throughput. The ability of the assay to process high numbers of lentiviral samples in a high throughput manner was proven by performing a virus stability study, demonstrating the effects of temperature, salt, and shear stress on LV infectivity.
format	article
author	Jennifer J Labisch G Philip Wiese Kalpana Barnes Franziska Bollmann Karl Pflanz
author_facet	Jennifer J Labisch G Philip Wiese Kalpana Barnes Franziska Bollmann Karl Pflanz
author_sort	Jennifer J Labisch
title	Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
title_short	Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
title_full	Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
title_fullStr	Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
title_full_unstemmed	Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
title_sort	infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/e96cb8de96504815a35df8cfd66633ec
work_keys_str_mv	AT jenniferjlabisch infectioustiterdeterminationoflentiviralvectorsusingatemporalimmunologicalrealtimeimagingapproach AT gphilipwiese infectioustiterdeterminationoflentiviralvectorsusingatemporalimmunologicalrealtimeimagingapproach AT kalpanabarnes infectioustiterdeterminationoflentiviralvectorsusingatemporalimmunologicalrealtimeimagingapproach AT franziskabollmann infectioustiterdeterminationoflentiviralvectorsusingatemporalimmunologicalrealtimeimagingapproach AT karlpflanz infectioustiterdeterminationoflentiviralvectorsusingatemporalimmunologicalrealtimeimagingapproach
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Infectious titer determination of lentiviral vectors using a temporal immunological real-time imaging approach.

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