• YAKHAK HOEJI
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• v.38 no.6
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• pp.715-720
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• 1994

${\alpha}2-macroglobulin$ $({\alpha}2-M)$ has been shown to have a variety of activities. One of those activities is the suppression of immune response. Characterization of the immunosuppressive factor secreted by a T cell hybridoma showed that ${\alpha}2-M$ was produced and secreted from the T cell hybridoma. ${\alpha}2-M$ was produced abundantly from the T cell hybridoma when cultured as ascites. The isolation and identification of the ${\alpha}2-M$ were studied using affinity chromatography and N-terminal amino acid sequencing. The extended observations were that the ${\alpha}2-M$ produced by the T cell hybridoma suppresses mixed lymphocyte reaction.

• Animal cells and systems
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• v.4 no.4
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• pp.381-388
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• 2000

Using a murine T cell hybridoma, activation-induced cell death (AICD) was studied. As an in vitro model system for the AICD, 1 cell hybridoma expressing TCR/CD3 complex was incubated onto the immobilized purified anti-CD3 antibody. The immobilized anti-CD3 antibody induced AICD effectively up to 40%. At 1-100 $\mu$M range of SNP, an exogenous source of nitric oxide (NO), the cell proliferation was not affected, but at 1 mM SNP, cell proliferation was significantly reduced. The AICD of T cell hybridoma was inhibited by exogenous NO at non-cytotoxic concentration, In the cells undergoing AICD, the expressions of caspase-3 and FasL were detected, but not iNOS. Similar result was recognized in the apoptosis induced by dexamethasone, an apoptosis-inducing agent. However, the conversion from the inactive form of caspase-3 (32 kDa) to the active form (17 kDa) was significantly reduced in the cells in AICD induced by anti-CD3 antibody, With the result of increased PARP cleavage in the cells, we propose that another PARP cleavage pathway not involving caspase-3 may function in the anti-CD3 antibody induced AICD in the T cell hybridoma.

• BMB Reports
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• v.30 no.6
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• pp.415-420
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• 1997

Although $CD4^+$ T cell responses to protein-derived antigen have well been understood, the epitopes recognized by hapten-specific $CD4^+$ T cells have not been fully defined. In this study, we characterized the response of a T cell hybridoma (5Di0.1B8) which is specific for a hapten. N-hydroxysuccinimidyl-4-azidobenzoate (HSAB) restricted by MHC class II $I-A^d$. Using three different antigen presenting cells (APCs) expressing $I-A^d$, the role of class II MHC proteins in haptenic antigen presentation and subsequent activation of 5D10.1B8 has been examined. Activation of 5D10.1B8 T cells by HSAB analogs was also performed. Our results show that each APC activated T cells differentially and that interleukin-2 (IL-2) augmented antigen-presenting ability of all the APCs, suggesting that increased expression of class II MHC protein by IL-2 played an important role in HSAB presentation and T cell activation. Finally, early T cell receptor-dependent signals induced by HSAB or its analogs were examined by phosphotyrosine immunoblot analysis, and showed that tyrosine phosphorylation level of a 18-20 kD protein increased upon stimulation.

• KSBB Journal
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• v.13 no.1
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• pp.108-113
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• 1998

The cell growth and amino acid metabolism of a hybridoma cell line in T-flasks, spinner flasks, and a 2L bioreactor were compared. Similar growth and metabolic behaviour were observed for spinner flask and bioreactor cultivations, while those in T-flasks differed significantly. Through a detailed analysis of nutrients and wastes, 7 amino acids were found to be consumed to a much higher extent than the rest of the amino acids. Supplementing the based medium with selected amino acids, glucose, and vitamines increased the cell density by 70%. The addition of vitamines was found to increase the metabolic rates of glucose and lactate.

• Journal of Microbiology and Biotechnology
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• v.2 no.2
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• pp.141-151
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• 1992

Physiological changes of the murine hybridoma cell line $S3H5/\gamma{2bA2}$ during adaptation to RPMI 1640 medium with 1%(v/v) fetal bovine serum were characterized in terms of cell growth, antibody production, morphology, and metabolic quotients. Cells adapted to 1% serum medium in T-flasks became sensitive to shear induced by mechanical agitation and required at least 5% serum in the medium or spent medium for cell growth in spinner flasks, while cells adapted to 10% serum medium in T-flasks could grow in 1% serum medium in spinner flasks. Consequently, long-term adaptation to low serum media may not give the expected growth enhancement. After adaptation to 1% serum medium, changes in cell morphology were observed. The cells in 10% serum medium were uniform and circular, while cells in 1% medium were irregularly shaped. The DNA contents, which were measured by flow cytometry, were almost constant among the cells in the range of 1% to 10%. Further, no significant changes in energy metabolism and specific monoclonal antibody production rate were observed among these cells.

• KSBB Journal
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• v.13 no.1
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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.

• Biotechnology and Bioprocess Engineering:BBE
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• v.2 no.2
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• pp.73-76
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• 1997

Biopolymer membrane was prepared using two oppositely charged natural biopolymer. The biopolymer membrane was used for the encapsulation of two hybridoma cell lines(ATCC CRL-1606, ATCC BH-8852) to produce monoclonal antibodies. In order to reduce the down stream steps, the pore 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$107cells/mL 3$\times$107cells/mL, and MAb concentrations of 506 $\mu\textrm{g}$/mL and 109$\mu\textrm{g}$/mL for encapsulated ATCC CRL-1606 and HB-8852, respectively. Two liter perfusion culture with encapsulated ATCC HB-8852 was performed to enhance the MAb production. The MAb production of the encapsulated hybridoma increased considerably comparing to the culture using silicone tubing for oxygen transfer.

• Animal cells and systems
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• v.2 no.3
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• pp.403-410
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• 1998

The Tcf-1 (T cell specific factor-1) is a transcription factor uniquely expressed in T-lineage cells. Its expression is developmentally regulated, which is high in the specific stage of immature thymocytes, but is much lower in mature T cells. We cloned the Tcf-1 gene by subtractive hybridization and found it to be highly expressed in the thymus compared to the mRNA level in the spleen as expected. Since apoptosis occurs enormously in the thymus, we were interested in whether Tcf-1 gene expression could be regulated by such a high level of apoptotic assault. By using T cell hybridoma 70.7 cells, we induced apoptosis by incubating cells with anti-CD3 antibody in vitro. After apoptosis induction, Tcf-1 mRNA level was found to be significantly reduced compared to normal cells. Since Tcf-1 is a transcription factor for the CD3-e gene, we tested how CD3-e expression is regulated in apoptotic cells. The surface level of CD3-e protein is also down-regulated after apoptosis induction. Such a down-modulation of CD3-e protein would reduce the TCR/CD3 complex on the cell surface, which would be an important regulator for T cell apoptosis.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.2
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• pp.110-117
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• 2000

Relationship between monoclonal antibody (MAb) productivity and growth rate, and effects of high cell density on MAb production rate increased with increasing specifis growth rate in both suspended and immobilized continuous cultures indicate a positively growth-associated relationship between MAb productivity and growth rate. moreover, the specific production rate was higher in the immobilized cell culture than that in suspended one at all dilution rates. In order to clarify these phenomana, MAb mPNA experession and cell cycle distribution were investigated in bacth cultures with immobilized cells and suspended cells. RT-PCR was used for observation of MAb mRNA expression and a two-color bromodeoxyuridine (BrdU)/propidium iodide (PI) flow cytometry method for determination of cell cycle distribution. The results revealed that MAb nRNA expression until dead phase, which was longer than in suspended cell. The cell cycle distribution patterns were observed almost the same for both immobilized and suspended cells. Such results may imply that a high cell density state has positive influence on the mRNA expression and on growth-associated Mab productivity of T0405 cells.

• Korean Journal of Veterinary Research
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• v.39 no.6
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• pp.1112-1118
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• 1999

Galectin-3 plays an important role in cell development, differentiation and cancer metastasis, including cell-cell/extracellular matrix (ECM) interactions and is supposed to have an effect of apoptosis on T-cells in thymic clonal selection. In this study, to know the effect of galectin-3 on thymocyte development, we used recombinant human galectin-3 (rHgal-3) from Escherichia coli, JM105, which was inserted with human gal-3 gene-transformed plasmid vector (prGal-3) to express human galectin-3. Expressed rHgal-3 was confirmed by western blot using the culture supernant of hybridoma (M3/38) producing monoclonal antibody to human galectin-3. Sepharose gel affinity chromatography was used to purify the expressed rHgal-3. Thymocytes and hepatocytes from 6-week-old male BALB/c mice were incubated with rHgal-3 and showed marked increase of apoptotic population on analysis using flow cytometry with 7-AAD in a dosedependent manner. However, rHgal-3 failed to induce apoptosis on hepatocytes. Interestingly, this apoptotic effect was not inhibited by lactose, a specific lectin domain inhibitor. From these results, we concluded that extrinsic -3 induces apoptosis on mouse thymocytes, and galectin-3 may have an apoptotic effect on T-cells in thymic clonal selection.