Production and purification of high-titer OrfV for preclinical studies in vaccinology and cancer therapy
| Autor: | Grant McFadden, Jessica A. Minott, Thomas M. McAusland, Byram W. Bridle, Jacob P. van Vloten, Sarah K. Wootton, Lisa A. Santry, Jim Petrik, Joelle C. Ingrao |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
Cancer therapy
OrfV QH426-470 Biology Virus 03 medical and health sciences 0302 clinical medicine Protocol Genetics medicine High titer gradient ultracentrifugation Molecular Biology 030304 developmental biology 0303 health sciences QH573-671 tangential flow filtration Cancer biology.organism_classification medicine.disease Virology 3. Good health Parapoxvirus ovis poxvirus 030220 oncology & carcinogenesis virus purification Parapoxvirus cancer therapy Molecular Medicine vaccinology Cytology |
| Zdroj: | Molecular Therapy: Methods & Clinical Development, Vol 23, Iss, Pp 434-447 (2021) Molecular Therapy. Methods & Clinical Development |
| ISSN: | 2329-0501 |
| Popis: | Poxviruses have been used extensively as vaccine vectors for human and veterinary medicine and have recently entered the clinical realm as immunotherapies for cancer. We present a comprehensive method for producing high-quality lots of the poxvirus Parapoxvirus ovis (OrfV) for use in preclinical models of vaccinology and cancer therapy. OrfV is produced using a permissive sheep skin-derived cell line and is released from infected cells by repeated freeze-thaw combined with sonication. We present two methods for isolation and purification of bulk virus. Isolated virus is concentrated to high titer using polyethylene glycol to produce the final in vivo-grade product. We also describe methods for quantifying OrfV infectious virions and determining genomic copy number to evaluate virus stocks. The methods herein will provide researchers with the ability to produce high-quality, high-titer OrfV for use in preclinical studies, and support the translation of OrfV-derived technologies into the clinic. Graphical abstract Methods for producing high-titer, high-purity OrfV, a poxvirus of the Parapoxvirus ovis genera and a promising oncolytic and vaccine vector, are described, as are methods for accurate titration of OrfV. These methods will provide researchers with the knowledge to produce in vivo-grade OrfV for preclinical studies. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|