An automated HIV-1 Env-pseudotyped virus production for global HIV vaccine trials
Infections with HIV still represent a major human health problem worldwide and a vaccine is the only long-term option to fight efficiently against this virus. Standardized assessments of HIV-specific immune responses in vaccine trials are essential for prioritizing vaccine candidates in preclinical...
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| Published in: | PloS one Vol. 7; no. 12; p. e51715 |
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| Main Authors: | , , , , , , , , , |
| Format: | Journal Article |
| Language: | English |
| Published: |
United States
Public Library of Science
27-12-2012
Public Library of Science (PLoS) |
| Subjects: | |
| Online Access: | Get full text |
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| Summary: | Infections with HIV still represent a major human health problem worldwide and a vaccine is the only long-term option to fight efficiently against this virus. Standardized assessments of HIV-specific immune responses in vaccine trials are essential for prioritizing vaccine candidates in preclinical and clinical stages of development. With respect to neutralizing antibodies, assays with HIV-1 Env-pseudotyped viruses are a high priority. To cover the increasing demands of HIV pseudoviruses, a complete cell culture and transfection automation system has been developed.
The automation system for HIV pseudovirus production comprises a modified Tecan-based Cellerity system. It covers an area of 5×3 meters and includes a robot platform, a cell counting machine, a CO(2) incubator for cell cultivation and a media refrigerator. The processes for cell handling, transfection and pseudovirus production have been implemented according to manual standard operating procedures and are controlled and scheduled autonomously by the system. The system is housed in a biosafety level II cabinet that guarantees protection of personnel, environment and the product. HIV pseudovirus stocks in a scale from 140 ml to 1000 ml have been produced on the automated system. Parallel manual production of HIV pseudoviruses and comparisons (bridging assays) confirmed that the automated produced pseudoviruses were of equivalent quality as those produced manually. In addition, the automated method was fully validated according to Good Clinical Laboratory Practice (GCLP) guidelines, including the validation parameters accuracy, precision, robustness and specificity.
An automated HIV pseudovirus production system has been successfully established. It allows the high quality production of HIV pseudoviruses under GCLP conditions. In its present form, the installed module enables the production of 1000 ml of virus-containing cell culture supernatant per week. Thus, this novel automation facilitates standardized large-scale productions of HIV pseudoviruses for ongoing and upcoming HIV vaccine trials. |
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| Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Competing Interests: The authors have declared that no competing interests exist. Conceived and designed the experiments: SK AS MS-K DAO. Performed the experiments: SK AS MF ASM. Analyzed the data: SK AS MS-K DAO. Contributed reagents/materials/analysis tools: DM. Wrote the paper: AM AS. Conceptional outline and participating by adding key points to the manuscript: AM MS-K DM HVB HZ. |
| ISSN: | 1932-6203 1932-6203 |
| DOI: | 10.1371/journal.pone.0051715 |