Evaluation of JEV and BVDV Clearance During the Purification of Recombinant HPV-16 L1 Virus-Like Particles.
- Author:
Jung Yun CHOI
1
;
Hye Sung JEONG
;
Young Lim KIM
;
Hwa Kyung SON
;
Byoung Guk KIM
;
Seung Rel RYU
;
Jin Ho SHIN
;
Sun Young BAEK
;
Hong Ki MIN
;
Sue Nie PARK
Author Information
1. Department of Biologics Evaluation, Korea Food and Drug Administration, 5 Nokbun-dong Eunpyung-gu, Seoul, 122-704, Korea. suenie@kfda.go.kr
- Publication Type:Original Article
- Keywords:
Recombinant HPV-16 L1 VLPs;
Viral clearance;
JEV;
BVDV
- MeSH:
Blotting, Western;
Capsid Proteins;
Cell Line;
Centrifugation;
Cesium;
Detergents;
Diarrhea;
Electrophoresis, Polyacrylamide Gel;
Encephalitis Virus, Japanese;
Human papillomavirus 16*;
Humans;
Insects;
Sf9 Cells;
Sonication;
Sucrose
- From:Journal of Bacteriology and Virology
2004;34(3):213-221
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
Insect cell-derived biotechnological products have a potential for viral contamination from cell line sources themselves or from adventitious introduction of virus during production. The objective of this study was to establish techniques for viral clearance validation of insect cell-derived recombinant human papillomavirus (HPV)-16 type L1 virus-like particles (VLPs) using Japanese encephalitis virus (JEV) and bovine viral diarrhea virus (BVDV) as relevant viruses. The downstream process for the production of recombinant HPV-16 L1 VLPs was sequentially carried out employing detergent lysis (NP-40/PBS), sonication, sucrose cushion centrifugation, and cesium chloride (CsCl) equilibrium density centrifugation. Recombinant HPV-16 L1 capsid protein (56 kD) expressed in Sf9 cell culture was clearly detected by SDS-PAGE and Western blotting analysis. Each purification step was evaluated to determine reduction factor for viral clearance by infectivity assay. In individual purification steps, detergent treatment (0.50% v/v, NP-40/PBS) and CsCl equilibrium density centrifugation were found to be effective in JEV and BVDV clearance. Overall cumulative reduction factors of JEV and BVDV infectivity titer for the purification procedure implemented in this study were 12.53 and 10.05 log TCID(50)/pool, respectively. The results suggest that the purification procedure employed in this study for the HPV-16 L1 VLPs produced from recombinant baculovirus-infected Sf9 cells will be effective over 10 log TCID(50)/pool reduction factor in the clearance of enveloped, adventitious viruses with a buoyant density lower than approximately 1.23 g/ml.
