A Four-Step Purification Process for Gag VLPs: From Culture Supernatant to High-Purity Lyophilized Particles


Posted: 2021-10-26 19:00:00
Vaccines (Basel) . 2021 Oct 9;9(10):1154. doi: 10.3390/vaccines9101154. Affiliations Expand Affiliations 1 Departament d'Enginyeria Química Biològica i Ambiental, Universitat Autònoma de Barcelona, 08193 Barcelona, Spain. 2 Department of Bioengineering, McGill University, Montreal, QC H3A 0E9, Canada. Item in Clipboard Irene González-Domínguez et al. Vaccines (Basel). 2021. Show details Display options Display options Format Vaccines (Basel) . 2021 Oct 9;9(10):1154. doi: 10.3390/vaccines9101154. Affiliations 1 Departament d'Enginyeria Química Biològica i Ambiental, Universitat Autònoma de Barcelona, 08193 Barcelona, Spain. 2 Department of Bioengineering, McGill University, Montreal, QC H3A 0E9, Canada. Item in Clipboard CiteDisplay options Display options Format Abstract 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 × 109 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. Keywords: EVs; Gag VLPs; chromatography; clarification; lyophilization; nanoparticle quantification; purification. Grant support

参考サイト PubMed: exsome



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