Application of aqueous two-phase micellar system to improve extraction of adenoviral particles from cell lysate
Biotechnol. appl. biochem
; 65(3): 381-389, 2018. ilus, tab
Article
in En
| SES-SP, SESSP-IDPCPROD, SES-SP
| ID: biblio-1060882
Responsible library:
BR79.1
Localization: BR79.1
ABSTRACT
Viral vectors are important in medical approaches, such as disease prevention and gene therapy, and their production depends on efficient prepurification steps. In the present study, an aqueous two-phase micellar system (ATPMS) was evaluated to extract human adenovirus type 5 particles from a cell lysate. Adenovirus was cultured in human embryonic kidney 293 (HEK-293) cells to a concentration of 1.4 × 1010 particles/mL. Cells were lysed, and the system formed by direct addition of Triton X-114 in a 23 full factorial design with center points. The systems were formed with Triton X-114 at a final concentration of 1.0, 6.0, and 11.0% (w/w), cell lysate pH of 6.0, 6.5, and 7.0, and incubation temperatures at 33, 35, and 37 °C. Adenovirus particles recovered from partition phases were measured by qPCR. The best system condition was with 11.0% (w/w) of Triton X-114, a cell lysate pH of 7.0, and an incubation temperature at 33 °C, yielding 3.51 × 1010 adenovirus particles/mL, which increased the initial adenovirus particles concentration by 2.3-fold, purifying it by 2.2-fold from the cell lysate, and removing cell debris. In conclusion, these results demonstrated that the use of an aqueous two-phase micellar system in the early steps of downstream processing could improve viral particle extraction from cultured cells while integrating clarification, concentration, and prepurification steps.
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Collection:
06-national
/
BR
Database:
SES-SP
/
SESSP-IDPCPROD
Main subject:
Water
/
Genetic Vectors
Limits:
Humans
Language:
En
Journal:
Biotechnol. appl. biochem
Year:
2018
Document type:
Article