Abstract
We have previously shown that hepatitis delta virus (HDV) virus-like particles (VLPs) can be assembled and secreted by hepatoma cells co-transduced by two recombinant baculoviruses (Bac-GD encoding L-HDAg and Bac-GS2 encoding HBsAg). In this study, we further explored the ability of assembly and secretion of VLP in different mammalian cells. The co-transduced BHK cells led to the formation of VLPs resembling authentic virions in size and appearance and the optimal dosage ratio of Bac-GS2 to Bac-GD for BHK was found to be 4. The production process was further transferred to a novel oscillating packed bed bioreactor, BelloCell500. The BHK cells cultured in BelloCell500 (1.5×109) were co-transduced at 27 °C by 125 ml Bac-GS2 and 25 ml Bac-GD, supplemented with 375 ml DPBS as surrounding solution. Using this protocol, the transduction efficiency (90%) and mean fluorescence intensity (FI) were high. Besides, the HDV VLPs were secreted into the medium as demonstrated by electron microscopy, among them about 46% were 37-nm particles. In order to simplify the purification process, the serum free medium (VP-SFM) was used, which resulted in similarly high transduction efficiency and mean FI, but lower VLP yield (only 33 % of that obtained from serum containing medium). The VLPs in the conditioned medium was concentrated by Stirred Cell with a membrane size of 300 kDa, and then purified by the combination of ion exchange and con A affinity chromatography.