Ventini-Monteiro Daniella C, Astray Renato M, Pereira Carlos A
Laboratório de Imunologia Viral, Instituto Butantan, CEP 05503-900, São Paulo, Brazil.
Methods Mol Biol. 2018;1674:87-94. doi: 10.1007/978-1-4939-7312-5_7.
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.
一次性固定床生物反应器(iCELLis nano)可用于培养非贴壁昆虫细胞,随后可回收这些细胞以进行扩大培养,或用于收获在蛋白质结构和生物学功能保存方面具有高质量的膜相关病毒糖蛋白。在此,我们描述了在iCELLis nano生物反应器中建立转基因黑腹果蝇Schneider 2(S2)细胞培养物以及通过酶联免疫吸附测定(ELISA)对合成的重组狂犬病病毒糖蛋白(rRVGP)进行定量的步骤。通过使用所描述的生产方案,可定期实现以下性能:总细胞数为1.7±0.6×1E10;细胞密度为2.4±0.8×1E7个细胞/毫升,每1E7个细胞合成1.2±0.9微克rRVGP;rRVGP总量为1.5±0.8毫克。