• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1997년도 춘계학술대회
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• pp.93-93
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• 1997

Analysis of protein is often frustrated by the inability to isolate large amounts of purified protein from a native source. To overcome this problem, fusion protein expression systems such as pGEX system have been widely used. Using pGEX system, the desired protein could be easily obtained in a large amount in E. coli, and then the fusion protein could be used for the study of the function of the given protein. To analyze and purify the GST fusion protein, anti-GST antibody could be used as one of the system of choice. However, the production and characterization of monoclonal anti-GST antibody has not been studied extensively yet. To produce monoclonal anti-GST antibody, GST was purified from E. coli transformed with pGEX-cs, one of the pGEX system and was used as an antigen. The monoclonal antibody was produced by fusion of the immunized spleen cells with SP2-0 myeloma cells. The antibody was characterized by ELISA, western blotting, etc. The monoclonal antibody produced in this study (mAb-GSTA) showed strong and specific immunoreactivity against not only GST but also GST-fusion proteins. Also, mAb-GSTA was successfully used for the immunoaffinity purification of the GST ${\beta}$-Rc.-third intracellular-loop fusion protein. The results of the present study suggest that mAb-GSTA may be used for the identification and purification of GST fusion proteins.

• 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.

• Journal of Microbiology
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• 제34권4호
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• pp.379-383
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• 1996

Cryptosporidium parvum causes a life-threatening diarrhea in acquired immunodeficiency syndrome (AIDS) patients. THe sporozoite stage of C. parvum has been known to be a target in treating cryptosporidiosis in AIDS patients as it is an extracellular stage. A sporozoite was ultrastructurally observed. It has a creascent shape with a rounded posterior end and a tapering body. The compact nucleus was located at the posterior end. A monoclonal antibody was produced, which recognized a 43 kDa of sporozoite antigens in a western blot analysis and showed the surface labeling in immunofluorescence.

• Biomolecules & Therapeutics
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• 제5권4호
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• pp.344-350
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• 1997

The purpose of the present study was to produce and characterize a monoclonal antibody against human ${\beta}_2$-adrenergic receptor. Male BALB/c mice were immunized with glutathione S-transferase (GST) fusion protein of the C-terminal portion of the human ${\beta}_2$-adrenergic receptor which was expressed in E.Coli. The immunized splenocytes were fused with myeloma SP2/0-Agl4 cells. The resulting hybridomas were screened for the production of a monoclonal antibody which can recognize human ${\beta}_2$-adrenergic receptor, and then subcloned by limiting dilution. The resulting monoclonal antibody was named as mAb$\beta$CO2. The mono-clonal antibody $\beta$CO2 was determined as IgM subtype and then purified by anti-mouse IgM-agarose affinity chromatography. The results of ELISA, Western blot, and immunocytochemistry showed that mAb$\beta$CO2 recognized human ${\beta}_2$-adrenergic receptor in the ${\beta}_2$-adrenergic receptor-GST fusion protein and human spider-moid carcinoma cell line A431 with highly specific immunoreactivity, The monoclonal antibody $\beta$CO2 may provide useful tools for the study of the $\beta$-adrenergic receptor of human and other species including rats.

• 한국미생물·생명공학회지
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• 제19권3호
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• pp.272-280
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• 1991

유가식 배양에서 배지첨가속도가 하이브리도마 세포의 증식과 항체생성속도에 미치는 영향을 조사하였다. 항체 비생성속도 $q_p$값이 변화는, 회분식 배양의 경우 배양 초기에 가장 높았다가 계속 감소하는 경향을 보였으나, 배양 48시간 이후부터 배지의 첨가가 시작되는 유가식 배양에서는 qp 값이 배지 첨가직후 부터 빠른 속도로 증가하기 시작해 배양이 끝날 때까지 높은 값으로 유지되었다. 그리고 배지의 비첨 가속도 $q_f$가 증가되면서 항체의 비생성속도 $q_p$도 민감하게 증가하는 경향을 보였다. 생성된 최종 항체농도 $C_{final}$은 배지 첨가속도에 비해 큰 영향을 받지 않고 거의 비슷한 값을 보였지만 평균적으로 회분식 배양보다는 그 값이 3배 이상 증가되어 최고 220$\mu g$/ml까지 향상되었다.

• KSBB Journal
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• 제13권1호
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• pp.13-19
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• 1998

Biopolymer membrane was prepared using two oppositely charged natural biopolymers. The biopolymer membrane was used for the encapsulation of two hybridoma cell lines(ATCC CRL-1606, ATCC HB-8852) to produce monoclonal antibodies. In order to reduce the down stream steps, the pre size of the membrane was controlled to retain the monoclonal antibodies in the capsules based on the diffusion experiments with standard proteins. T-flask culture showed cell densities of 8$\times$107 cells/mL and 3$\times$107 cells/mL, and MAb concentrations of 506$\mu$g/mL and 109$\mu$g/mL for encapsulated ATCC CRL-1606 and HB-8852, respectively. Two liter perfusion cultures with encapsulated ATCC HB-8852 were performed to enhance the MAb production. The MAb production of the encapsulated hybridoma increased considerably comparing to the culture using silicon tubing for oxygen transfer.

• 대한수의학회지
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• 제44권4호
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• pp.625-635
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• 2004

Trichnuris suis does not excrete eggs during larval stage as well as in particular adult stage, It is impossible to diagnose by use of fecal examination method in those periods. Therefore, serological diagnostic method can be very useful for those stages. In order to produce monoclonal antibody, specific somatic and secretory-excretory (SE) antigens of T. suis were identified and analyzed by SDS-PAGE and Western blot. Monoclonal antibody-producing hybridoma cells were cloned, which were made of popliteal lymph node of BALB/c mice immunized with a 45 kDa somatic antigen of T. suis. Five clones (1B9, 2C4, n2C5, 2D7 and 2D8) showing strong responses to T. suis antigens were selected and the isotype identified. All monoclonal antibodies were IgG1 isotype and the light chains were k chain. Established monoclonal antibodies reacted specifically to somatic and SE antigens of T. suis and did not cross-reacted to antigens of ascaris suum, trichuris vulpis, or Trichinella spiralis. The sensitivity of somatic and SE antigens against these monoclonal antibodies were significant (p<0.01) associated with those of positive and negative sera.

• 한국동물학회지
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• 제35권1호
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• pp.29-36
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• 1992

The present study describes the production of monoclonal antibodies against cultured chick myoblast to pursue critical proteins in muscle cell fusion. Among a panel of monoclonal antibodies, three, Mll-3H 13, Mll-3Hl8 and Mll-3H35 were inhibited movblast fusion. A single 101-kDa antigen reactive with monoclonal antibody Mll-3H35 was detected by radioimmu-noprecipitation or by immunoblotting. During the course of myogenesis, the level of the protein remarkably decreased as the cells there differentiated. These results suggest that the protein platys a direct role in the process of myoblast fusion mechanism.

• 대한수의학회지
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• 제30권4호
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• pp.511-513
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• 1990

$11{\alpha}$-hydroxyprogesterone hemisuccinate-BSA를 항원으로 하여 항원량을 $50{\mu}g/head$ 및 $100{\mu}g/head$의 2 group(세마리씩)으로 나누어 BALB/c mouse에 면역첩종한 결과 후자에서 항체가의 상송이 확인되었다. 이와 동시에 항원($20{\mu}g$)과 adjuvant의 비율을 1:9로 하여 장기 접종한 결과는 $100{\mu}g$투여시 보다 항체가가 낮았다. 항체가가 확인된 clone의 culture에 의해 progesterone 단일크론 hybridoma를 생산해 이의 supernatant에 대한 분석을 실시한 결과 immunoglobulin class는 IgM이었다. progesterone 이외의 다른 steroids와의 교차반응은 매우 낮았다.

• KSBB Journal
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• 제11권3호
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• pp.329-339
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• 1996

The effects of ammonium ion on cell growth kinetics, monoclonal antibody productivity, and cell metabolism of hybridoma cells were investigated. The mouse-mouse hybridoma cell line VlIIH-8 producing mouse IgG2a was used as a model system. Ammonium ion showed an inhibitory effect on cell growth and monoclonal antibody production. New immobilized adsorbents were developed for the reduction of the inhibitory effect of ammonium ion. The ammonium ion selective zeolite, Phillipsite-Gismondine was entrapped in calcium alginate bead or in dialysis membrane and applied to the hybridoma cell culture system for the in situ removal of ammonium ion from culture media. The effects of ammonium the both serum supplemented and serum free media on the cell growth were studied by applying immobilized adsorbents of calcium alginate bead type. The results demonstrated a substantial enhancement in cell growth. Applying immobilized adsorbents of dialysis membrane type to serum supplemented media also resulted in the stimulation of cell growth, cell viability and monoclonal antibody production.