Dataset Information

Optimization of lentiviral vector production for scale-up in fixed-bed bioreactor.

ABSTRACT: Lentiviral vectors (LVs) are promising tools for gene therapy. However, scaling up the production methods of LVs in order to produce high-quality vectors for clinical purposes has proven to be difficult. In this article, we present a scalable and efficient method to produce LVs with transient transfection of adherent 293T cells in a fixed-bed bioreactor. The disposable iCELLis bioreactors are scalable with a large three-dimensional (3D) growth area range between 0.53 and 500?m², an integrated perfusion system, and a controllable environment for production. In this study, iCELLis Nano (2.67-4?m²) was used for optimizing production parameters for scale-up. Transfections were first done using traditional calcium phosphate method, but in later runs polyethylenimine was found to be more reliable and easier to use. For scalable LV production, perfusion rate control by measuring cell metabolite concentrations in the bioreactor leads to higher productivity and reduced costs. Optimization of cell seeding density for targeted cell concentration during transfection, use of low compaction fixed-bed and lowering the culture pH have a positive effect on LV productivity. These results show for the first time that iCELLis bioreactor is scalable from bench level to clinical scale LV production.

SUBMITTER: Valkama AJ

PROVIDER: S-EPMC5817386 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Optimization of lentiviral vector production for scale-up in fixed-bed bioreactor.

Valkama A J AJ Leinonen H M HM Lipponen E M EM Turkki V V Malinen J J Heikura T T Ylä-Herttuala S S Lesch H P HP

Gene therapy 20171005 1

Lentiviral vectors (LVs) are promising tools for gene therapy. However, scaling up the production methods of LVs in order to produce high-quality vectors for clinical purposes has proven to be difficult. In this article, we present a scalable and efficient method to produce LVs with transient transfection of adherent 293T cells in a fixed-bed bioreactor. The disposable iCELLis bioreactors are scalable with a large three-dimensional (3D) growth area range between 0.53 and 500 m<sup>2</sup>, an in ...[more]

PMID: 29345252

Similar Datasets

Project description:Rice wine, well known for its unique flavor, rich nutritional value, and health benefits, has potential for extensive market development. Rhizopus and Aspergillus are among several microorganisms used in rice wine brewing and are crucial for determining rice wine quality. The strains were isolated via Rose Bengal and starch as a combined separation medium, followed by oenological property and sensory evaluation screening. The strain exhibiting the best performance can be screened using the traditional rice wine Qu. The strains YM-8, YM-10, and YM-16, which exhibited strong saccharification and fermentation performance along with good flavor and taste, were obtained from traditional rice wine Qu. Based on ITS genetic sequence analysis, the YM-8, YM-10, and YM-16 strains were identified as Rhizopus microsporus, Rhizopus arrhizus, and Aspergillus oryzae. The optimum growth temperature of each of the three strains was 30°C, 32°C, and 30°C, and the optimum initial pH was 6.0, 6.5, and 6.5, respectively. The activities of α-amylase, glucoamylase, and protease of YM-16 were highest at 220.23±1.88, 1,269.04±30.32, and 175.16±1.81 U/g, respectively. The amino acid content of rice wine fermented in a 20-L bioreactor with the three mold strains was higher than that of the control group, except for arginine, which was significantly lower than that of the control group. The total amino acid content and the total content of each type of amino acid were ranked as YM-16 > YM-8 > YM-10 > control group, and the amino acid content varied greatly among the strains. The control group had a higher content, whereas YM-8 and YM-16 had lower contents of volatile aroma components than the control group and had the basic flavor substances needed for rice wine, which is conducive to the formation of rice wine aroma. This selected strain, YM-16, has strong saccharification and fermentation ability, is a rich enzyme system, and improves the flavor of rice wine, thereby demonstrating its suitability as a production strain for brewing.

Project description:Studies investigating the feasibility of new, or improved, biotechnologies, such as wastewater treatment digesters, inevitably start with laboratory-scale trials. However, it is rarely determined whether laboratory-scale results reflect full-scale performance or microbial ecology. The Expanded Granular Sludge Bed (EGSB) bioreactor, which is a high-rate anaerobic digester configuration, was used as a model to address that knowledge gap in this study. Two laboratory-scale idealizations of the EGSB-a one-dimensional and a three- dimensional scale-down of a full-scale design-were built and operated in triplicate under near-identical conditions to a full-scale EGSB. The laboratory-scale bioreactors were seeded using biomass obtained from the full-scale bioreactor, and, spent water from the distillation of whisky from maize was applied as substrate at both scales. Over 70 days, bioreactor performance, microbial ecology, and microbial community physiology were monitored at various depths in the sludge-beds using 16S rRNA gene sequencing (V4 region), specific methanogenic activity (SMA) assays, and a range of physical and chemical monitoring methods. SMA assays indicated dominance of the hydrogenotrophic pathway at full-scale whilst a more balanced activity profile developed during the laboratory-scale trials. At each scale, Methanobacterium was the dominant methanogenic genus present. Bioreactor performance overall was better at laboratory-scale than full-scale. We observed that bioreactor design at laboratory-scale significantly influenced spatial distribution of microbial community physiology and taxonomy in the bioreactor sludge-bed, with 1-D bioreactor types promoting stratification of each. In the 1-D laboratory bioreactors, increased abundance of Firmicutes was associated with both granule position in the sludge bed and increased activity against acetate and ethanol as substrates. We further observed that stratification in the sludge-bed in 1-D laboratory-scale bioreactors was associated with increased richness in the underlying microbial community at species (OTU) level and improved overall performance.

Dataset Information

Optimization of lentiviral vector production for scale-up in fixed-bed bioreactor.

Publications

Optimization of lentiviral vector production for scale-up in fixed-bed bioreactor.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets