• Journal of Microbiology and Biotechnology
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• 제31권3호
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• pp.349-357
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• 2021

Monoclonal antibodies are widely used as diagnostic reagents and for therapeutic purposes, and their demand is increasing extensively. To produce these proteins in sufficient quantities for commercial use, it is necessary to raise the output by scaling up the production processes. This review describes recent trends in high-density cell culture systems established for monoclonal antibody production that are excellent methods to scale up from the lab-scale cell culture. Among the reactors, hollow fiber bioreactors contribute to a major part of high-density cell culture as they can provide a tremendous amount of surface area in a small volume for cell growth. As an alternative to hollow fiber reactors, a novel disposable bioreactor has been developed, which consists of a polymer-based supermacroporous material, cryogel, as a matrix for cell growth. Packed bed systems and disposable wave bioreactors have also been introduced for high cell density culture. These developments in high-density cell culture systems have led to the monoclonal antibody production in an economically favourable manner and made monoclonal antibodies one of the dominant therapeutic and diagnostic proteins in biopharmaceutical industry.

• Biotechnology and Bioprocess Engineering:BBE
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• 제5권2호
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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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• 제1권2호
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• pp.126-129
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• 1991

In order to produce ${\gamma}-linolenic$ acid by Mucor sp. KCTC 8405P. the fungus was cultivated in fed-batch culture with two phases. i.e., growth in yeast-like form and induction to hyphal growth by pH shift of the culture medium during cultivation. The synchronous growth of the fungus into the appropriate sizes was important for the high density cell culture of this dimorphic fungus. Dissolved oxygen concentration in the medium did not affect degree of unsaturation of fatty acids and ${\gamma}-linolenic$ acid content. Under the culture conditions applied in this experiment. the fungus was found to produce 100 g/l dry mycelia containing 40% of the lipids, where ${\gamma}-linolenic$ acid comprised about 9% of the total extractable fatty acids.

• Journal of Microbiology and Biotechnology
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• 제19권12호
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• pp.1603-1611
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• 2009

High-cell-density cultivation of yeast was investigated using the agricultural waste products corn steep liquor (CSL) and molasses. The Saccharomyces cerevisiae KV-25 cell mass was significantly dependent on the ratio between C and N sources. The concentrations of molasses and CSL in the culture medium were statistically optimized at 10.25% (v/v) and 16.87% (v/v), respectively, by response surface methodology (RSM). Batch culture in a 5-l stirred tank reactor using the optimized medium resulted in a cell mass production of 36.5 g/l. In the fed-batch culture, the feed phase was preceded by a batch phase using the optimized medium, and a very high dried-cell-mass yield of 187.63 g/l was successfully attained by feeding a mixture of 20% (v/v) molasses and 80% (v/v) CSL at a rate of 22 ml/h. In this system, the production of cell mass depended mainly on the agitation speed, the composition of the feed medium, and the glucose level in the medium, but only slightly on the aeration rate.

• ALGAE
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• 제33권1호
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• pp.127-141
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• 2018

Low efficiency in microalgal biomass production was largely attributed to the low density of algal cell cultures. Though mutations that reduced the level of chlorophyll or pigment content increased efficiency of photon usage and thus the cell-culture density under high-illumination growth conditions (e.g., >$500{\mu}mol\;photon\;m^{-2}\;s^{-1}$), it was unclear whether algae could increase cell-culture density under low-illumination conditions (e.g., ${\sim}50{\mu}mol\;photon\;m^{-2}\;s^{-1}$). To address this question, we performed forward genetic screening in Chlamydomonas reinhardtii. A pool of >1,000 insertional mutants was constructed and subjected to continuous subcultures in shaking flasks under low-illumination conditions. Complexity of restriction fragment length polymorphism (RFLP) pattern in cultures indicated the degree of heterogeneity of mutant populations. We showed that the levels of RFLP complexity decreased when cycles of subculture increased, suggesting that cultures were gradually populated by high cell-culture density (HCD) strains. Analysis of the 3 isolated HCD mutants after 30 cycles of subcultures confirmed that their maximal biomass production was 50-100% higher than that of wild type under low-illumination. Furthermore, levels of chlorophyll content in HCD mutant strains were similar to that of wild type. Inverse polymerase chain reaction analysis identified the locus of insertion in two of three HCD strains. Molecular and transcriptomic analyses suggested that two HCD mutants were a result of the gain-of-function phenotype, both linking to the abnormality of mitochondrial functions. Taken together, our results demonstrate that HCD strains can be obtained through continuous subcultures under low illumination conditions.

• Journal of Microbiology and Biotechnology
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• 제18권5호
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• pp.918-925
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• 2008

Controlling the light energy and major nutrients is important for high cell density culture of cyanobacterial cells. The growth phase of Anabaena variabilis can be divided into an exponential growth phase and a deceleration phase. In this study, the cell growth in the deceleration phase showed a linear growth pattern. Both the period of the exponential growth phase and the average cell growth rate in the deceleration phase increased by controlling the light intensity. To control the light intensity, the specific irradiation rate was maintained above $10\;{\mu}mol/s/g$ dry cell by increasing the incident light intensity stepwise. The final cell density increased by controlling the nutrient supply. For the control of the nutrient supply, nitrate, phosphate, and sulfate were intermittently added based on the growth yield, along with the combined control of light intensity and nutrient concentration. Under these control conditions, both final cell concentration and cell productivity increased, to 8.2 g/l and 1.9 g/l/day, respectively.

• Biotechnology and Bioprocess Engineering:BBE
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• 제8권6호
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• pp.349-353
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• 2003

The physiological characteristics of cultures of very high cell mass (e.g. 10g cell mass/L), termed“ultrahigh cell density cultures”is reviewed. A close relationship was found between the length of the optical path (OP) in flat-plate reactors and the optimal cell density of the culture as well as its areal (g m$\^$-2/ day$\^$-1/) productivity. Cell-growth inhibition (GI) unfolds as culture density surpasses a certain threshold. If it is constantly relieved, a 1.0cm OP reactor could produce ca. 50% more than reactors with longer OP, e.g. 5 or 10cm. This unique effect, discovered by Hu et al. [3], is explained in terms of the relationships between the frequency of the light-dark cycle (L-D cycle), cells undergo in their travel between the light and dark volumes in the reactor, and the turnover time of the photosynthetic center (PC). In long OP reactors (5cm and above) the L-D cycle time may be orders of magnitude longer than the PC turnover time, resulting in a light regime in which the cells are exposed along the L-D cycle, to long, wasteful dark periods. In contrast, in reactors with an OP of ca. 1.0 cm, the L-D cycle frequency approaches the PC turnover time resulting in a significant reduction of the wasteful dark exposure time, thereby inducing a surge in photosynthetic efficiency. Presently, the major difficulty in mass cultivation of ultrahigh-density culture (UHDC) concerns cell growth inhibition in the culture, the exact nature of which is awaiting detailed investigation.

• KSBB Journal
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• 제9권2호
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• pp.230-237
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• 1994

회전식 여과기는 일종의 분리기로서 동물세포의 연속배양에 이용되어 높은 세포농도와 그와 비례한 높은 생산성을 가져다 준다. 회전식 여과기를 이용한 세포배양은 여러 인자들에 의해 세포농도의 변화가 결정되는데 이를 수학적으로 modeling하고 수치 모사와 sensitivity analysis를 통하여 조사하였다. 고농도 배양시 암모니아의 축적은 세포 성장을 크게 둔화시키고 최대세포농도도 따라서 낮게 된다. 운전 인자 중 세포유치율은 세포 성장속도와 최대세포농도의 크기에 가장 큰 영향을 끼침이 밝혀졌다. 비배지공급속도는 세포농도의 변화에 거의 영향을 끼치지 않으며 배지의 연속식공급과 계단식공급은 세포성장에 큰 차이를 보이지 않는다.

• KSBB Journal
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• 제14권2호
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• pp.160-166
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• 1999

본 연구에서는 ($\beta$-carotene 생산균주인 미세조류 Dunaliella bardawil을 사용하여 batch flask에서 미세조류의 고농도세포에 관한 최적배양조건(미량원소, pH, agitation speed, nitrate, phosphate, carbon source)을 확립하고자 하였다. 미량원소는 5X 배지에서 교반하였을 때 비생장속도는 $0.0l3hr^{-l}$와 세포농도는 $4.9{\times}10^6$ cells/mL로서 IX. 3X, lOX 배지에서 배양한 것보다. 약 46%, 18%, 69% 높은 세포수율을 얻었으며 세포배양시 교반한 경우, pH는 80에서 최대 세포농도를 얻었다. 초기 nitrate ($KNO_3$)와 phosphate($KH_2PO_4$)의 영향을 조사한 결과 미세조류 생장에 중요한 영양분으로서 질소원의 주입은 매우 효과적임을 확인하였다. 또힌 탄소원으로서 250mM의 $NaHCO_3$와 $CO_2$ 가스를 동시에 사용한 배양조건이 500mM $NaHCO_3$만을 탄소원으로 사용한 실험에 비하여 32% 증가된 세포농도를 나타내었다. light는 white light의 경우 blue light보다 세포생장에 적합하였다. 질소원을 이용한 유기배양시 2회의 nitrate주입만으로써 배양 198hr에 $8.955{\times}10^6$cell/mL의 고농도의 세포를 얻었다.

• KSBB Journal
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• 제6권3호
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• pp.231-240
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• 1991

Recently, developments of large scale and high density cell culture methods have been the objects of many researches, because the demand of various pharmaceutical products produced by animal cell culture has been rapidly increasing. The cell culture equipment should have the requirements such as sufficient oxygen transfer and mixing, low shear stress and surface tension, and small foaming. In order to develop a proper bioreactor meeting these requirements simultaneously, a perfluorocarbon having high solubility of oxygen was sprayed into the medium as an oxygen carrier instead of air. Also, a new impeller was developed and combined together with the perfluorocarbon spraying system so as to design a new bioreartor for cell cultivation. The new impeller had better characteristics of mixing and oxygen transfer than the paddle and cell-lift impellers based on the same, shear rate. But, it was observed that the volumetric oxygen transfer coefficient of the new bioreactor decreased with increasing cell density during E. coli fermentation.