• 생명과학회지
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• 제24권12호
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• pp.1301-1307
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• 2014

지방유래줄기세포(adipose-derived stem cell; ADSC)는 조직재생을 위한 탁월한 수단으로 인정되고 있으나 세포증식속도가 느려 ADSC의 배양용 배지에는 대게 fetal bovine serum (FBS)이 첨가된다. FBS는 세포에 다양한 영양분을 공급하지만 세포의 기능에 영향을 미칠 수 있는 미 동정 물질도 많이 함유하고 있다. FBS에 의한 예상밖의 영향과 동물유래물질의 오염을 방지하기 위해 ADSC의 무혈청 배양법에 관한 연구가 광범위하게 이루어지고 있다. 본 연구에서는 ADSC세포에 SV40의 T항원 유전자를 도입하여 증식속도를 향상시킨 ADSC-T세포주의 효율적인 무혈청 배양법을 확립하기 위해 ADSC-T의 세포증식에 미치는 아미노산복합체, 비타민 복합체 및 여러가지 영양분 혼합물(B27)의 영향을 검토하였다. 그 결과, ADSC-T세포를 DMEM/F12 무혈청 배지에 현탁하여 plate에 주입하였을 때는 증식하지 않았으며 아미노산, 비타민 및 B27 영양소복합체는 증식촉진효과를 나타내지 않았다. 그러나 ADSC-T세포를 유혈청 DMEM배지로 24시간 배양 후 DMEM/F12 무혈청 배지로 교체하여 배양했을 때는 세포가 증식하였으며 이때, 비타민 복합체와 B27 영양소복합체는 증식촉진효과를 나타내었다. 또한 Stem pro 배지를 이용하면 ADSC-T의 무혈청 부유배양이 가능한 것으로 나타났다. ADSC-T세포는 분자량 70 kDa 부근의 단백질을 다량으로 분비하였으며, 성장인자 중에서 insulin-like growth factor (IGF)와 fibroblast growth factor basic (FGF basic)는 유혈청 배양보다 무혈청 배양에서 더 많이 분비되었다.

• KSBB Journal
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• 제5권1호
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• pp.87-94
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• 1990

본 논문에서는 10% fetal bovine serum(FBS)를 첨가한 RPMI 1640배지에 비하여 간염바이러스 표면항원에 대한 단일클론항체 생산에 있어서 효과적은 무혈청 배지에 낮은 온도의 혈청을 첨가하여 하이브리도마_2_c3.1 세포에서 그 효과를 조사하였다, 세포성장관 단일클론항체 생산을 증가시키기 위하여 기본 무혈청배지와 RPMI 1640 배지성분들의 농도를 균형있게 강화시킨 배지를 2:1(v / v)로 혼합하여 농축배지를 조성하고, 이 배지에 2mg / ml 인혈청 알부민 $5\;{\mu\textrm{g}}\;/\;ml$ insulin, $5\;{\mu\textrm{g}}\;/\;ml$ tran-sferrin, $10\;{\mu\textrm{g}}\;/\;ml$ monoethanolmine 등의 몇가지 무혈청 첨가물들을 첨가하였다. 이 농축 배지에 fetal bovine serum(FBS)과 supplemented bovine calf serum(sBCS)을 첨가하였을때의 세포성장과 단일클론항체 생산을 비교하여 FBS농도를 변화하여 세포성장과 단일클론항체생산의 증가를 시도하였다. 0.5%를 농축배지에 첨가함으로써 세포성장과 단일클론항체생산의 증가를 보았다. 최대 세포농도는 $3.06{\times}10^6$ cells / ml이었으며, 이때 생산된 단일클론항체는 10% FBS배지의 $43.0\;{\mu\textrm{g}}\;/\;ml$ 무혈청배지의 $50\;{\mu\textrm{g}}\;/\;ml$보다 3배이상 높은 $159.7\;{\mu\textrm{g}}\;/\;ml$이 생산되었다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2005년도 생물공학의 동향(XVI)
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• pp.108-108
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• 2005

• 한국수정란이식학회지
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• 제24권4호
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• pp.281-287
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• 2009

As a simple and economical method for in vitro produced embryos, we have used BSA instead of serum for the production and embryo transfer of Hanwoo in vitro fertilized (IVF) embryos and obtained the following results: 1) When using serum (FBS; fetal bovine serum) or BSA-containing culture media as the initial culture media for immature oocytes, it is regarded as inappropriate to add only BSA to the culture solutions from maturation of the immature oocytes to development stage culture, but serum still needs be added though there is no significant difference in the concentration, with a change from 5% to 10%. 2) The results of culturing IVF embryos after development (4 cell stage) in the Medium199 solutions containing BSA instead of serum (FBS) showed that 0.3% BSA concentration is not optimal and 0.5% or higher BSA concentration has no significant difference among 0.5%, 0.7%, 1% and 2% (p > 0.05). 3) The post-freezing survival ratio after development in 5% FBS-Medium199 showed that 1% BSA concentration of the culture solution is the most suitable in the BSA concentrations of 0.3% (51%), 0.5% (67%), 0.7% (69%), 1% (77%) and 2% (75%). 4) The pregnancy rates of the transplanted fresh(not frozen) blastocyst had no significant concentration dependency (p > 0.5), and the average pregnancy rate was 63.8%. 14% of overweight calves were found among the calves given birth to by the transfer of IVF blastocysts cultured in the serum-added culture solution, but none was found in the experimental groups in which BSA was added instead of serum.

• Journal of Microbiology
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• 제42권1호
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• pp.25-31
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• 2004

Sf9 cells have obvious advantages for the conventional production technology of vaccine. They are useful tools for high concentration and large-scale cultures. Sf9 cells were grown to maximal concentration, 8${\times}$l0$\^$6/ cells/$m\ell$ in a 500$m\ell$ spinner flask, with a doubling time at the exponentially growing phase of 24.5 hours, using serum-free media. To explore the ability of Sf9 cells to be infected by the Japanese encephalitis (JE) virus Beijing-l strain, Sf9 cells were infected with the virus. By 4-5 days post-infection, 10-15 % of the Sf9 cells showed cytopathic effect (CPE), from granularity to the formation of syncytia and multinucleated giant cells continuously observed over a period of 35 days. Positive fluorescent reactions were detected in 30-40% of cells infected with the JE virus Beijing-l strain, and the uninfected Sf9 cells were completely negative. Virus particles, propagated in Sf9 and Vero cells, were concentrated by sedimentation on 40% trehalose cushions by ultracentrifugation, and showed identical patterns of viral morphogenesis. Complete virus particles, 40 to 50 nm in diameter, were observed, and JE virus envelope (E) proteins, at 53 kDa, were found in the western blot analysis to the anti-JE virus E protein monoclonal antibody and reacted as a magenta band in the same position to the glycoprotein staining. To evaluate whether the infectious virus was produced in Sf9 cells inoculated with the JE virus Beijing-l stain, Sf9 cells were inoculated with the virus, and sample harvested every 5 days. The titers of the JE virus Beijing-l strain rose from 1.0${\times}$l0$\^$5/ to 1.5${\times}$l0$\^$6/ pfu/$m\ell$. The infected Sf9 cells could be subcultured in serum-free medium, with no change in the plaque sizes formed by the JE virus Beijing-l strain in the plaque assay. It is suggested that the ability of the JE virus Beijing-l strain to infect Sf9 cells in serum-free media will provide a useful insect cell system, where the JE virus replication, cytopathogenicity and vaccine immunogen can be studied.

• KSBB Journal
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• 제21권2호
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• pp.110-117
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• 2006

일반적인 세포주의 동결보존은 혈청이 첨가된 배양배지에 10% DMSO를 첨가하여 실행하게 된다. 그러나 무혈청 환경에서 배양되는 세포주를 이용하여 생물의약품을 생산하는 공정에 사용되는 세포주의 경우는 교차오염 방지를 위해 동결보존 역시 혈청을 제거한 상태에서 실시되어야 한다. 본 실험은 무혈청 동결보존이 CHO 세포에 어떠한 영향을 미치는지 알아보기 위하여 실시되었다. 우선, 무혈청 동결보존에서 세포 생존율을 높이기 위해 투과성 및 비투과성 첨가제를 배지에 첨가하는 방법으로 동결보존 및 해동하여 생존율을 측정하였다. 그 결과, 10% DMSO와 0.03 M raffinose를 동시에 첨가한 경우에 76%의 생존율을 확보할 수 있었지만 기존의 혈청을 이용하는 동결보존에는 미치지 못하였다. 두 번째로 무혈청 배지의 동결보존 능력을 알아보기 위해 시판 중인 무혈청 배지와 무혈청 동결보존제를 사용하여 동결 보존 후의 생존율을 비교한 결과, 무혈청 배지를 이용한 실험에서는 혈청 배지를 이용한 동결보존과 유사한 95% 이상의 생존율을 확인할 수 있었다. 이는 무혈청 동결보존제의 동결보존 능력보다 우수한 결과이다. 마지막으로 무혈청 배지를 이용한 장기간 동결보존에서 CHO 세포의 안정성을 확인하기 위하여 무혈청 배지로 동결보존된 CHO 세포를 3개월 단위로 해동하여 생존율 및 성장 회복율을 측정하고, real-time RT-PCR을 통해 삽입된 CHO DHFR 유전자의 안정성을 평가하였다. 혈청배지와 비교할 때, 생존율과 성장 회복율, 유전자 안정성 측면에서 모두 동일한 결과를 보여, 무혈청 배지를 이용한 18개월 이상의 동결보존이 안정적임을 검증할 수 있었다. 결과적으로 생물의약품 공정에서 무혈청 배지를 이용한 동결보존이 혈청을 이용한 동결보존을 대체할 수 있을 것으로 생각된다.

• Parasites, Hosts and Diseases
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• 제36권2호
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• pp.99-108
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• 1998

베네수엘라분선충의 충란, 자충 띤 성충을 장기간 보존할 수 있는 방안을 마련하기 위하여 여러 가지 배양환경에 대한 연구를 수헝하였던 바, 다음과 같은 결과를 얻었다. 분변 내의 충란은 $4^{\circ}C$ 에서 25일간, 실온에서는 15일간 생존하였으며, 분변에서 분리한 충란은 $4^{\circ}C$의 생리적 식염수에 서 24시간 생존하였으나, 상실배기의 충란은 건조한 공기에 민감하여 12시간 내에 환성을 잃었다. 감염형 자충을 polyvinylbag에 넣어 $20^{\circ}C$에 보관하였을 경우, 분변 물질이 첨가된 0.12% 영양배 지에서는 45일, 영양배지는 28일간 생존하였다 한편, 영양배지에서 배양된 감염 자충은 상수에서 는 32일간, 멸균 식염수에서는 22일간 생존하였다. 인공감염시켜 얻은 성충은 $37^{\circ}C$에서 9일간, 9:1배지 (10% 쥐 혈청을 첨가)와 혈청을 첨가하지 않은 영양배지를 매일 교환하여 주어도 4일동안 밖에 생존하지 못하였다. 성충 암컷은 실험관 내에서 산란하고, 자충으로 부화되었지만 혈청을 첨가한 배지에서도 감염자충으로 발육하지 못하였다.

• KSBB Journal
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• 제5권4호
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• pp.365-371
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• 1990

To improve the growth of mouse hybridoma 2c3.1 secreting anti-Hepatitis B surface antigen monoclonal antibody (anti-HBsAg MAb), we had constructed an enriched medium and observed the effects of fetal bovine serum and serum-free supplements including human serum albumin, 'insulin and transferrin', and monoethanolamine. For further enhancement of growth, the concentrations of two major energy sources, glucose and glutamine, were strengthened with various ratios in the enriched medium. Maximum cell growth and monoclonal antibody production obtained in various ratios of glucose/glutamine with an inoculation concentration of 2$\times$105 cells/ml were 0.73$\times$106-4.62$\times$106 cells/ml and 65.1-422.6 $\mu\textrm{g}$/ml, respectively. Glutamine was round to be a major energy source and a limiting nutrient in comparison to glucose for 2c3.1 cell cultivation in enriched media with low serum.

• 대한한방내과학회지
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• 제19권2호
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• pp.392-410
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• 1998

In order to investigate the effect of Alismatis Lhizoma on hyperlipidemia, experimental studies were performed on hyperlipidemia rats. Hyperlipidemia model (controll group) was induced by 1% cholesterol fed-diet for 8 weeks. Sample I group fed with 1% cholesterol and 4% Alismatis Lhizoma diet for 8 weeks. Sample II group fed with 1% cholesterol and 8% Alismatis Lhizoma diet for 8 weeks. The contents of serum total cholesterol, triglyceride, free fatty acid, phospholipid, HDL-cholesterol and LDL-cholesterol were measured, and fat accumulation in liver and the change of elastic and collagenous fiber in aortic wall were observed. The results were summurized as follows ; 1. The content of total cholesterol in the serum compared with control group tended to be decreased in sample group, and then sample I group showed a significant value. 2. The content of triglyceride in the serum compared with control group tended to be decreased in sample group, and then sample I group showed a significant value. 3. The content of free fat acid in the serum compared with control group tended to be decreased in sample group, but did not show a significance. 4. The content of phospholipid in the serum compared with control group tended to be decreased in sample group, and then sample I group showed a significant value. 5. The content of HDL-cholesterol in the serum compared with control group tended to be increased in sample group, and then sample II group showed a significant value. 6. The content of LDL-cholesterol in the serum compared with control group tended to be decreased in sample group, and then sample I group showed a significant value. 7. The lipophagy in liver compared with control group tended to be repressed in sample group. 8. The change of elastic and collagenous fiber lesion in tunica media of aortic wall, compared with control group tended to be repressed in sample group. According to the above results, it is assumed that Alismatis Lhizoma has a valid effect on hyperlipidemia. And yet, it needs to make further researches that sample I group showed more significant value than sample II group.

• 대한한방내과학회지
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• 제19권2호
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• pp.347-366
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• 1998

In order to investigate the effect of Lycii Radicis Cortex on hyperlipidemia, experimental studies were performed on hyperlipidemia rats. Hyperlipidemia model (controll group) was induced by 1% cholesterol fed-diet for 8 weeks. Sample I group fed with 1% cholesterol and 4% Lycii Radicis Cortex diet for 8 weeks. Sample II group fed with 1% cholesterol and 8% Lycii Radicis Cortex diet for 8 weeks. The contents of serum total cholesterol, triglyceride, free fatty acid, phospholipid, HDL-cholesterol and LDL-cholesterol were measured, and fat accumulation in liver and the change of elastic and collagenous fiber in aortic wall were observed. The results were summurized as follows ; 1. The content of total cholesterol in the serum compared with control group tended to be decreased in sample group, but did not show a significance. 2. The content of triglyceride in the serum compared with control group tended to be decreased in sample group, and then sample II group showed a significant value. 3. The content of free fat acid in the serum compared with control group tended to be decreased in sample group, and then sample II group showed a significant value. 4. The content of phospholipid in the serum compared with control group tended to be decreased in sample group, but did not show a significance. 5. The content of HDL-cholesterol in the serum compared with control group tended to be increased in sample group, and then sample I group showed a significant value. 6. The content of LDL-cholesterol in the serum compared with control group tended to be decreased in sample group, and then sample I group showed a significant value. 7. The lipophagy in liver compared with control group tended to be decreased in sample group. 8. The change of elastic and collagenous fiber lesion in tunica media of aortic wall, compared with control group tended to be decreased in sample group. According to the above results, it is assumed that Lycii Radicis Cortex has a valid effect on hyperlipidemia. Therefore, it seems to be applicable to the diseases related to hyperlipidemia.