• Korean Journal of Veterinary Research
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• v.26 no.2
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• pp.259-264
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• 1986

Microcarrier culture technics are widely used for the massive production of the vertebrate animal cells. In this study, attempt was made to establish the microcarrier cell culture system using Cytodex. Various factors affecting the growth of cells on microcarrier culture were also discussed. In conclusion, the yield of cells in microcarrier culture was several times greater than those in roller bottle and flask culture methods, based on the volume of culture media.

• Microbiology and Biotechnology Letters
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• v.17 no.3
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• pp.231-235
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• 1989

Possibility of using microcarriers for the growth of a transformed human embryonic kidney cell line 293 was investigated. The cell grew well in a static culture such as T-flasks with medium of DME/F12 (3:1) mixture supplemented with 5% FBS, but it was most difficult to make the cells grow on microcarriers mainly due to the low attachment efficiency and poor spreading at initial stage of the culture. Consequently, 30-50% of the cells were lost upon inoculation into microcarrier suspension and significant fraction of the mirrocarrier became bald. The medium supplemented with the concentrated conditioned medium by hepatoma cell line HpG2 supported the active growth of the cells on microcarrier and the cells showed a very healthy and well spreading morphology. It was probable that some spreading and attachment factors of HpG2 conditioned medium were effective for 293 cells.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.2
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• pp.123-129
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• 2000

A high cell density culture system for the anchorage dependent CHO cells was developed based on the combination of in removal of ammonium ion and microcarrier culture system, and semi-fed-batch feeding of glucose and glutamine was employed to the developed culture system. The glass bead was selected as an optimum microcarrier in terms of cell growth. An ammonium ion selective zeolite, Phillipsite-Gismondine, was packed in a dialysis menium ion. The semi-fed-batch operation was employer to the novel culture system for the high density cell culture, and the results showed the cell growth was improved by 32% and tPA productivity by 250%.

• Journal of Microbiology and Biotechnology
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• v.23 no.9
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• pp.1308-1321
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• 2013

The emergence of microcarrier technology has brought a renewed interest in anchorage-dependent cell culture for high-yield processes. Well-known in vaccine production, microcarrier culture also has potential for application in other fields. In this work, two types of microcarriers were evaluated for small-scale monoclonal antibody (mAb) production by CHO-K1 cells. Cultures (5 ml) of microporous Cytodex 3 and macroporous CultiSpher-S carriers were performed in vented conical tubes and subsequently scaled-up (20 ml) to shake-flasks, testing combinations of different culture conditions (cell concentration, microcarrier concentration, rocking methodology, rocking speed, and initial culture volume). Culture performance was evaluated by considering the mAb production and cell growth at the phases of initial adhesion and proliferation. The best culture performances were obtained with Cytodex 3, regarding cell proliferation (average $1.85{\pm}0.11{\times}10^6$ cells/ml against $0.60{\pm}0.08{\times}10^6$ cells/ml for CultiSpher-S), mAb production ($2.04{\pm}0.41{\mu}g/ml$ against $0.99{\pm}0.35{\mu}g/ml$ for CultiSpher-S), and culture longevity (30 days against 10-15 days for CultiSpher-S), probably due to the collagen-coated dextran matrix that potentiates adhesion and prevents detachment. The culture conditions of greater influence were rocking mechanism (Cytodex 3, pulse followed by continuous) and initial cell concentration (CultiSpher-S, $4{\times}10^5$ cells/ml). Microcarriers proved to be a viable and favorable alternative to standard adherent and suspended cultures for mAb production by CHO-K1 cells, with simple operation, easy scale-up, and significantly higher levels of mAb production. However, variations of microcarrier culture performance in different vessels reiterate the need for optimization at each step of the scale-up process.

• KSBB Journal
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• v.13 no.1
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• pp.83-89
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• 1998

Using a cellulose macroporous microcarrier, HeLa cells were cultivated in 100mL spinner flask(Bellco Co., USA) and confluent cell laden microcarriers were subcultured by bead-to-bead cell transfer method. In macroporous mcirocarrier-HeLa system viable suspended cells played an important role in bead-to-bead cell transfer and that could be increased by use of RPMI-1640, a calcium-ion-reduced-media and high speed agitation. Successful bead-to-bead cell transfers were performed continuously three time in spinner flask. We applied this technique to produce recombinant Vaccinia virus which express $\beta$-galactosidase. Recombinant protein yield of bead-to-bead transferred culture was comparable to conventional microcarrier cultures that were inoculated by cells detached from T-flask. Although trypsinization is a useful method for subculturing microcarriers in some cases, that process adds quality control problem and handling steps to large scale cell production. There fore, bead-to-bead cell transfer technique offers another convenient and efficient scale-up method for continuous microcarrier cultures.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.2
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• pp.118-122
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• 2000

A comlementary call line CR2 is curretly used to propagte the Disabled Infectious Single Cycle Herpes Simplex Virus Typee2 (DISC HSV-2) on a small Iaboratory scale upto 15 L. These cultures are initiated by passaging the cells from roller bottle cultures. Whilst this is suitable for the laboratory scale it is totally impractical for use in seeding an industrial manufacturing scaled version of the culture. It is paramount to have a robust system for passaging cells from a small microcarrierier culture system to a larger one by a serial subculturing regime. Here we report on the successes we have had in our laboratory in scaling up out production system for the DISC HSV-2 from small 1-L cultures to a 50-L vessel with the maintenance of the viral productivity. Ease of use, reproducibility and the need to minimise overall production time were factors which were taken into consideration whils developing our procedures. We were aware of the need to keep a production train simple and as short as possible as this was the amall scale study for an envisaged manufacturing process.

• KSBB Journal
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• v.11 no.2
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• pp.254-261
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• 1996

Attenuated varicella-zoster virus (VZV) was propagated in human lung fibroblast (HLF) cells Among media tested in this work, DMEM was the best medium for the growth of HLF cells. Because HLF was a normal finite cell line, cell growth late was dependent on the age of HLF cells. When the population doubling level (PDL) was higher than 46, apoptosis of HLF cells started and cell growth rate decreased. The optimum temperature for the cell growth and virus propagation in the T-flask culture was $37^{\circ}C$. In a microcarrier culture system in which Cytodex-3 was used for the VZV propagation in spinner vessels, the yield of plaque forming cells was lower than that in the T-flask culture. The relatively high shear environment near microcarriers was thought to cause the low yield of VZV in the microcarrier culture system.

• KSBB Journal
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• v.8 no.5
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• pp.465-472
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• 1993

The comparison of the capabilities of cell growth of four different kinds of commercially available microcarriers was carried out by culturing anchorage-dependent animal cells, Vero-6, in a spinner flask. Using 3 g/l of Cytodex 3, the maximum final cell density was about $1.4{\times}10^6$ cells/ml and increased up to $2.0{\times}10^6$ cells/ml by increasing microcarrier concentration up to 5 g/l. The macroporous collagen microcarriers, VX-100, informatrix, and Cultispher-G showed the final cell concentration of $4{\times}10^6$ cells/ml, $2.1{\times}10^6$ cells/ml, and $3.2{\times}10^6$ cells/ml, respectively at the microcarrier concentration of 5g/1. According to this result, VX-100 showed better cell growth than informatrix and cultispher-G and also showed about 2 fold increase in final cell density comparing to Cytodex 3 solid bead. When the intermittent bead-to-bead transfer technique was introduced in the culture using Cytodex 3 bead and cultispher-G, the result was very successful and the cells grew out very well. The recovered cells by dissolving collagen microcarrier using collagenase enzyme were mostly viable and grew out very well on the surface of the fresh microcarriers.

• KSBB Journal
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• v.13 no.3
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• pp.295-302
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• 1998

Human Erythropoietin (EPO) gene is cloned in quail fibrosarcoma cell, QT35. Because molecular weight of EPO is similar to that of serum albumin, cell culture with serum containing medium makes purification of EPO very difficult. Using fractional factorial study, we have developed serum free medium for the recombinant QT35 cell lines, QT N4D4 and QT SY-IMP, which have cloned EPO with glutamine synthetase (GS) gene amplification system and with puromycin selective marker, respectively. Among the seven frequently used medium components, fibronectin, BSA, and EGF were the most important for EPO production. However, sufficient fibronectin supplement to the medium did not make any good attachment of QT35 to culture plate over 3 days. Therefore, to maximize EPO production, we attempted a medium-shift at confluence from serum containing medium to serum free medium(QT SFM6). Using the medium-shift protocol with QT SFM6, nearly the same productivity of EPO was achieved comparing with that without medium-shift. This result was true in both QT35 cell lines in three types of culture, i.e. T flask, microcarrier and roller bottle cultures.