A four-step purification process for gag vlps : from culture supernatant to high-purity lyophilized particles
González-Domínguez, Irene (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Lorenzo, Elianet (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Bernier, Alice (McGill University. Department of Bioengineering)
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)
Kamen, Amine (McGill University. Department of Bioengineering)

Data:	2021
Resum:	Gag-based virus-like particles (VLPs) have high potential as scaffolds for the development of chimeric vaccines and delivery strategies. The production of purified preparations that can be preserved independently from cold chains is highly desirable to facilitate distribution and access worldwide. In this work, a nimble purification has been developed, facilitating the production of Gag VLPs. Suspension-adapted HEK 293 cells cultured in chemically defined cell culture media were used to produce the VLPs. A four-step downstream process (DSP) consisting of membrane filtration, ion-exchange chromatography, polishing, and lyophilization was developed. The purification of VLPs from other contaminants such as host cell proteins (HCP), double-stranded DNA, or extracellular vesicles (EVs) was confirmed after their DSP. A concentration of 2. 2 ± 0. 8 × 10 VLPs/mL in the lyophilized samples was obtained after its storage at room temperature for two months. Morphology and structural integrity of purified VLPs was assessed by cryo-TEM and NTA. Likewise, the purification methodologies proposed here could be easily scaled up and applied to purify similar enveloped viruses and vesicles.
Ajuts:	Ministerio de Educación, Cultura y Deporte FPU16/02555
Drets:	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.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Gag VLPs ; Purification ; Larification ; ChCromatography ; Lyophilization ; EVs ; Nanoparticle quantification
Publicat a:	Vaccines (Basel), Vol. 9, Issue 10 (October 2021) , art. 1154, ISSN 2076-393X

DOI: 10.3390/vaccines9101154
PMID: 34696262

19 p, 4.2 MB

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