Abstract
A single-use fixed-bed bioreactor (iCELLis nano) can be used for cultivating non adherent insect cells, which can be then recovered for scaling up or for harvesting a membrane-associated viral glycoprotein with high quality in terms of preserved protein structure and biological function. Here, we describe the procedures for establishing genetically modified Drosophila melanogaster Schneider 2 (S2) cell cultures in the iCELLis nano bioreactor and for quantifying by ELISA the recombinant rabies virus glycoprotein (rRVGP) synthesized. By using the described protocol of production, the following performance can be regularly achieved: 1.7 ± 0.6 × 1E10 total cells; 2.4 ± 0.8 × 1E7 cells/mL and 1.2 ± 0.9 μg of rRVGP/1E7 cells; 1.5 ± 0.8 mg of total rRVGP.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Loeffelholz C, Kaiser SC, Kraume M, Eibl R, Eibl D (2014) Dynamic single-use bioreactors used in modern liter-and m3 – scale biotechnological processes: engineering characteristics and scaling up. Adv Biochem Eng Biotechnol 138:1–44
Ventini DC, Astray RM, Lemos MAN, Jorge SAC, Riquelme CC, Suazo CAT, Tonso A, Pereira CA (2010) Recombinant rabies virus glycoprotein synthesis in bioreactor by transfected Drosophila melanogaster S2 cells carrying a constitutive or an inducible promoter. J Biotechnol 146:169–172
Ventini-Monteiro D, Dubois S, Astray RM, Castillo J, Pereira CA (2015) Insect cell entrapment, growth and recovering using a single-use fixed-bed bioreactor. Scaling up and recombinant protein production. J Biotechnol 216:110–115
Astray RM, Jorge SAC, Pereira CA (2017) Rabies vaccine development by expression of recombinant viral glycoprotein. Arch Virol 162:323–332. (in press)
Moraes AM, Jorge SAC, Astray RM, Suazo CAT, Calderón Riquelme CE, Augusto EFP, Tonso A, Pamboukian MM, Piccoli RAM, Barral MF, Pereira CA (2012) Drosophila melanogaster S2 cells for expression of heterologous genes: from gene cloning to bioprocess development. Biotechnol Adv 30:613–628
Astray RM, Augusto E, Yokomizo AY, Pereira CA (2008) Analytical approach for extraction of recombinant membrane viral glycoprotein from stably transfected Drosophila melanogaster cells. Biotechnol J 3:98–103
van Wezel AL (1967) Growth of cell-strains and primary cells on microcarriers in homogenous culture. Nature 216:64–65
Perrin P, Lafon M, Sureau P (1996) Enzyme linked immuno-sorbent assay (ELISA) for the determination of glycoprotein content of rabies vaccines. In: Meslin FX, Kaplan MM, Koprowski H (eds) Laboratory techniques in rabies. WHO, Geneva, pp 383–388
Acknowledgments
This work was supported in part by Artelis, ATMI LifeSciences, and PALL Life Sciences, and by a grant of CNPq (402439/2013-9). Carlos A Pereira is a recipient of CNPq 1A research fellowship.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Science+Business Media LLC
About this protocol
Cite this protocol
Ventini-Monteiro, D.C., Astray, R.M., Pereira, C.A. (2018). Production of Recombinant Rabies Virus Glycoprotein by Insect Cells in a Single-Use Fixed-Bed Bioreactor. In: Picanço-Castro, V., Swiech, K. (eds) Recombinant Glycoprotein Production. Methods in Molecular Biology, vol 1674. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7312-5_7
Download citation
DOI: https://doi.org/10.1007/978-1-4939-7312-5_7
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-7311-8
Online ISBN: 978-1-4939-7312-5
eBook Packages: Springer Protocols