Large-Scale Microcarrier Culture of Chinese Perch Brain Cell for Viral Vaccine Production in a Stirred Bioreactor

Abstract

Mandarin fish (Siniperca chuatsi) is one of the important cultured fish species in China. Infectious spleen and kidney necrosis virus (ISKNV) and Siniperca Chuatsi rhabdovirus (SCRV) have hindered the development of mandarin fish farming industry. Vaccination is the most effective method for control of viral diseases, however viral vaccine production requires the large-scale culture of cells. Herein, a suspension culture system of Chinese perch brain cell (CPB) was developed on Cytodex 1 microcarrier in a stirred bioreactor. Firstly, CPB cells were cultured using Cytodex 1 microcarrier in 125 mL stirring flasks. With the optimum operational parameters, CPB cells grew well, distributed uniformly, and could fully cover the microcarriers. Then, CPB cells were digested with trypsin and expanded step-by-step with different expansion ratios from the 125 mL stirring bottle to a 500 mL stirring bottle, and finally to a 3-L bioreactor. Results showed that with an expansion ratio of 1:3, we achieved a high cell density level (2.25 × 106 cells/mL) with an efficient use of the microcarriers, which also confirmed the data obtained from the 125 mL stirring flask. Moreover, obvious cytopathic effects (CPE) were observed in the suspended CPB cells post-infection with ISKNV and SCRV. This study provided a large-scale culture system of CPB cells for virus vaccine production.

Seed Train Optimization in Microcarrier-Based Cell Culture Post In Situ Cell Detachment through Scale-Down Hybrid Modeling

“…Conversely, a higher cells/bead ratio led to a higher final cell density within the same culture duration. The findings align with data generated in previous studies [ 19 , 21 , 48 , 49 ]. For example, Souza et al (2005) conducted a Design of Experiments (DoE) involving three parameters: agitation rate, cells/bead ratio, and FBS concentration.…”

“…Mainly, an additional control system is required to promote intermittent stirring [ 46 ]. Intermittent stirring is frequently recommended to increase contact time at the time of cell colonization on MCs [ 24 , 47 , 48 , 49 ]. For instance, Lue et al (2021) carried out experiments to assess the most effective stirring method in spinner flasks [ 49 ].…”

“…Intermittent stirring is frequently recommended to increase contact time at the time of cell colonization on MCs [ 24 , 47 , 48 , 49 ]. For instance, Lue et al (2021) carried out experiments to assess the most effective stirring method in spinner flasks [ 49 ]. They discovered that extending the duration of intermittent stirring led to better cell attachment, with the optimal condition identified as 3 min of stirring, followed by 42 min of standing over 5 h. However, it is worth noting that this observation was made only in small-scale experiments and has yet to be confirmed in larger scale settings.…”

Evaluation of Polymeric Particles for Modular Tissue Cultures in Developmental Engineering

“…In addition, the scaffolds with other forms such as membranes [ 15 ], sponges [ 16 ] and woven fabrics [ 17 ] are also fabricated according to the cell types and the targeted tissues. For the gradual layer-by-layer reconstruction of large and more functional 3D tissues with the equivalent sizes (scale: millimeter to meter) and structures of the target tissues, it is also necessary to manufacture sufficient various modular tissues via culturing different types of cells on the suitable modular scaffolds at large scale, which is comparable to the current microcarrier-based cell expansion systems [ 18 , 19 , 20 , 21 ].…”

The Stability and Efficency of CPB Cells Were Acclimated for Virus Proliferation

“…On the basis of the CPB cells line, our laboratory constructed an ISKNV and SCRV bivalent inactivated vaccine [ 16 ], and screened a SCRV potential live vaccine candidate for Chinese perch against SCRV disease [ 17 ]. For scale production of CPB cells for ISKNV and SCRV vaccines, we developed a suspension culture system of CPB cells on cytodex 1 microcarrier in 3-L bioreactor [ 18 ]. The stability, growth rate, and virus sensitivity of cells directly influence vaccine production scales, production costs, and competitiveness [ 19 , 20 , 21 ].…”