Transient expression of rabies virus G-glycoprotein using BHK-21 cells cultured in suspension
2015
Fernández-Núñez, Eutimio Gustavo | de Rezende, Alexandre Gonçalves | Puglia, Ana Lia Pradella | Leme, Jaci | Boldorini, Vera Lucia Lopes | Caricati, Celso Pereira | Tonso, Aldo
OBJECTIVE: To assess the expression of rabies virus G-glycoprotein (RVGP) expression using Semliki Forest virus as a vector in combination with BHK-21 cells cultured in suspension. RESULTS: A multilevel factorial design was used to quantify effects of temperature (33–37 °C), fresh medium addition after the viral adsorption step (100–200 % with respect to the initial cell suspension volume before infection) and harvest time (8–40 h) on RVGP production. Experimental runs were performed in 24-well cell culture plates at a multiplicity of infection (MOI) of 16. An additional experiment in spinner-flask was performed at MOI of 9, using the optimal conditions determined in cell culture plates. Values for temperature, fresh medium addition and harvest time of 33 °C, 100 % and 16 h, respectively, ensured the optimal RVGP production in culture plates. The volumetric yield (239 ng ml⁻¹) in these conditions was higher than that reported previously for adherent cell culture. In spinner-flasks, the volumetric yield was improved (559 ng ml⁻¹). CONCLUSION: These results establish the basis for designing bioprocess to produce RVGP.
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