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Vectored vaccines
Published in Amine Kamen, Laura Cervera, Bioprocessing of Viral Vaccines, 2023
Zeyu Yang, Kumar Subramaniam, Amine Kamen
Cultivation of mammalian cells has been realized using various technologies including roller bottles, microcarriers, bioreactor suspension cultures, etc. Suspension cell culture remains the most effective method for the production of AdV vectors at large scale especially when compared to processes using adherent cells. Additionally, with a homogeneous concentration of nutrients, metabolites, and cellular environment, the suspension culture is easy to monitor and scale-up to control process robustness and critical quality attributes of the AdV vectors. As the main cell line for production of AdV, HEK-293 cells have been adapted to suspension culture (293S). They were further adapted to serum-free medium 293SF, enhancing the scalability, batch-to-batch reproducibility, and regulatory approval [48,49]. Suspension cultures of HEK-293 in stirred tank bioreactors are projected to scale up to 10,000L, with yields for unpurified culture in the range of 109–1010 VP/ml [44,50]. Another important cell line PER.C6 has been successfully used in GMP manufacturing processes, growing to high cell densities in serum-free suspension culture, and can be used to produce AdV vectors in a similar fashion [50].
Amphiphilic Systems as Biomaterials Based on Chitin, Chitosan, and Their Derivatives
Published in Severian Dumitriu, Valentin Popa, Polymeric Biomaterials, 2020
Cationic methoxy poly(ethyleneglycol) (mPEG)-polyethylenimine (PE)-chitosan was synthesized77 and it showed a good DNA condensation capability, exhibiting negligible toxicity. The transfection of human embryonic kidney 293 (HEK 293) cells proved that mPEG-PEI-chitosan/VRaft-1 plasmid has little toxicity on the growth and gene expression of cells. Using this copolymer as transfection agent, the conversion of ω-6 to ω-3 fatty acids was successfully realized. Based on these superior properties, mPEG-PEI-chitosan should be a promising non-viral gene carrier for biomedical and biological applications.
A Review of Tubeless Microfluidic Devices
Published in Eric Lagally, Krzysztof Iniewski, Microfluidics and Nanotechnology, 2017
Pedro J. Resto, David J. Beebe, Justin C. Williams
Another clever example of passive pumping for automated cell manipulation was demonstrated by Ju et al. in their work using passive pumping in cell programmable assay (CPA) chips.28 The authors created CPA chips for culturing cells and automating the process of staining using surface tension passive pumping (Figure 9.31). The system was tested using human embryonic kidney (HEK) 293 cells. The rationale for this work was that interfacing robotics and culture control systems require equipment or methods that can make regular use of the device in small biology laboratories challenging. Therefore, they developed these chips to increase the throughput of common laboratory procedures without requiring the machines and equipment needed for most high-throughput approaches—for example, the high-throughput method proposed by Puccinelli et al. using ALHs.
Advances of engineered extracellular vesicles-based therapeutics strategy
Published in Science and Technology of Advanced Materials, 2022
Hiroaki Komuro, Shakhlo Aminova, Katherine Lauro, Masako Harada
HEK293 cell-derived EVs showed minimal toxicologically and immunogenic effects in mice as well as autologous-derived EVs in terms of functionality [104,105]. Some therapeutic agents produced by HEK293 cells have been approved by the US Food and Drug Administration (FDA) or European Medicines Agency (EMA) [106]. To enhance the therapeutic efficacy of EVs from HEK293 cells, cell engineering would be a requirement to load EVs with therapeutic cargo and target them to a specific tissue type. HEK293 cells can easily be manipulated with transfected plasmids or nucleic acids with targeting molecules. In addition, the fast growth and ease of culture of these cells allow for them to be more easily mass produced. EVs derived from several cell sources all exert different biological contents, functions, and biodistribution [107]. Exploiting the unique properties of EVs derived from different cell types will be an important factor for future treatments, including their large-scale generation and drug delivery efficiency. Therefore, in-depth studies are required for the development of EV-based therapies.
The bioactivity of titanium-cuttlefish bone-derived hydroxyapatite composites sintered at low temperature
Published in Powder Metallurgy, 2020
Anamarija Rogina, Ivona Košić, Maja Antunović, Marica Ivanković, Hrvoje Ivanković
Meanwhile, Hek293 cells were seeded in a 96-well plate (Sarsted) in triplicates at a density of 3 × 103 cells/200 μL and grown in Dulbecco’s modified Eagle medium with 4500 mg L−1 glucose (DMEM-high glucose, Lonza) containing 10% fetal bovine serum (FBS, Gibco), 100 U mL−1 penicillin and 100 mg mL−1 streptomycin (Sigma–Aldrich). After two days of culture, medium was replaced with powder extracts and incubated for next three days in 5% CO2 humidified atmosphere at 37°C. Following specified incubation periods (48 and 72 h), extracts were aspirated and 40 μL of MTT (Sigma–Aldrich) was added to each well at a concentration of 0.5 mg mL−1. After 4 h of incubation at 37°C, MTT was aspirated and formazan crystals were dissolved by adding 170 μL of DMSO to each well and incubated for 1 h at room temperature. Then, 100 μL of each solution was transferred into clean 96-well plate to measure the absorbance on a microplate reader (Promega) at 570 nm. The cell viability was calculated with respect to the non-treated cells. The cell morphology was analysed for each time point using Zeiss AxioVision inverted microscope.
Tricarbonylrhenium(I) complexes with the N,6-dimethylpyridine-2-carbothioamide ligand: combined experimental and calculation studies
Published in Journal of Coordination Chemistry, 2018
Krzysztof Lyczko, Monika Lyczko, Sylwia Meczynska-Wielgosz, Marcin Kruszewski, Józef Mieczkowski
For biological studies the following materials were used: RPMI-1640 medium, L-glutamine, cisplatin, phosphate-buffered saline (PBS), dimethylsulfoxide (DMSO) and 3-(4,5-dimetyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) were all purchased from Sigma Aldrich, eagle’s minimum essential medium (EMEM) was obtained from American Type Culture Collection (ATCC, Rockville, MD) and fetal calf serum (FCS) from Biological Industries (Israel). The human ovarian carcinoma A2780 cell line and its cisplatin resistant derivative, A2780cis, were purchased from Sigma Aldrich. Human cervical adenocarcinoma cells (HeLa) and human embryonic kidney cell line (Hek-293) were purchased from the ATCC. A2780 and HeLa cells were cultured in RPMI-1640 medium supplemented with 10% FCS and 2 mM L-glutamine. A2780cis cells were cultured in the same medium, but 1 μM cisplatin was added every 2–3 passages to maintain cisplatin resistance. Hek-293 cells were cultured in EMEM medium supplemented with 10% FCS. All cells were incubated in a 5% CO2 atmosphere at 37 °C.