Quality Assessment of Virus-Like Particles at Single Particle Level : A Comparative Study
González-Domínguez, Irene (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Puente-Massaguer, Eduard (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Cervera Gracia, Laura (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Gòdia, Francesc (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)

Date:	2020
Abstract:	Virus-like particles (VLPs) have emerged as a powerful scaffold for antigen presentation and delivery strategies. Compared to single protein-based therapeutics, quality assessment requires a higher degree of refinement due to the structure of VLPs and their similar properties to extracellular vesicles (EVs). Advances in the field of nanotechnology with single particle and high-resolution analysis techniques provide appealing approaches to VLP characterization. In this study, six different biophysical methods have been assessed for the characterization of HIV-1-based VLPs produced in mammalian and insect cell platforms. Sample preparation and equipment set-up were optimized for the six strategies evaluated. Electron Microscopy (EM) disclosed the presence of several types of EVs within VLP preparations and cryogenic transmission electron microscopy (cryo-TEM) resulted in the best technique to resolve the VLP ultrastructure. The use of super-resolution fluorescence microscopy (SRFM), nanoparticle tracking analysis (NTA) and flow virometry enabled the high throughput quantification of VLPs. Interestingly, differences in the determination of nanoparticle concentration were observed between techniques. Moreover, NTA and flow virometry allowed the quantification of both EVs and VLPs within the same experiment while analyzing particle size distribution (PSD), simultaneously. These results provide new insights into the use of different analytical tools to monitor the production of nanoparticle-based biologicals and their associated contaminants.
Grants:	Ministerio de Educación, Cultura y Deporte FPU16/02555 Ministerio de Educación, Cultura y Deporte FPU15/03577 Agència de Gestió d'Ajuts Universitaris i de Recerca 2017/SGR-898
Rights:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Language:	Anglès
Document:	Article ; recerca ; Versió publicada
Subject:	VLP ; Viral quantification ; NTA ; Flow virometry ; SRFM ; Cryo-TEM ; SEM
Published in:	Viruses, Vol. 12, Issue 2 (February 2020) , art. 223, ISSN 1999-4915

DOI: 10.3390/v12020223
PMID: 32079288

24 p, 8.2 MB

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Articles > Research articles
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