Hybridoma cells display an increase in antibody productivity following exposure to hypertonic conditions. However, the underlying mechanism is not well understood. In the present study, we hypothesize that the nuclear...Hybridoma cells display an increase in antibody productivity following exposure to hypertonic conditions. However, the underlying mechanism is not well understood. In the present study, we hypothesize that the nuclear factor of activated T cells 5 （NFAT5）/tonicity enhancer binding protein （TonEBP） functions to increase the antibody productivity of hybridoma cells. NFAT5 is an osmosensitive mammalian transcription factor. However, its ubiquitous expression in various organs that are not bathed in hypertonic milieu suggests that NFAT5 may also regulate cell growth and function under isotonic conditions. In this study, we examined the expression of NFAT5 in hybridoma cells by Western blot analysis, and found that it increased significantly in hypertonic medium. To further define the function of NFAT5 in hybridoma cells, RNA interference technique was used to downregulate the expression of NFAT5 in SGB-8 cells （a hybridoma cell line）. In isotonic medium, antibody productivity ofhybridoma cells was reduced by downregulation of NFAT5 while cell proliferation was not influenced. The results presented here demonstrate that NFAT5 not only plays an important role in osmotic stress response pathway in hybridoma cells but also is essential for optimal antibody productivity.展开更多
It has been developed in this laboratory that a serum-free medium, designated asDMI,is adaptive for long-term culture,freezing and resurgence in liquid nitrogen,and cloningof hybridoma and parental myelomas as well as...It has been developed in this laboratory that a serum-free medium, designated asDMI,is adaptive for long-term culture,freezing and resurgence in liquid nitrogen,and cloningof hybridoma and parental myelomas as well as for cell fusions.In this report,it is describedthat the myeloma cells grown in DMI for more than 3 months maintained their biological charac-teristics such as induction of aseites and subcutaneous somatic tumor in BALB/c mice and toler-ance toward 8-azaguanine,ete..However,the ability of the tumor cells to attach to glass wallwas decreased.The selecting assay of these cells in HAT medium(medium containing hypoxan-thine,aminopterin and thymidine)showed that the death time in DMI was similar to that in theconventional 15％ newborn calf serum-supplemented medium(RPS15).The hybridoma cellsadapted in DMI secrete monoclonal antibodies in quantities comparable to those produced inRPS15.展开更多
To get the hybridoma cell lines secreting anti-thyrotropin monoclonal antibodies with high affinity and specificity. Methods: BALB/c mice were immunized with extract of human pituitaries. The spleen cells of one immun...To get the hybridoma cell lines secreting anti-thyrotropin monoclonal antibodies with high affinity and specificity. Methods: BALB/c mice were immunized with extract of human pituitaries. The spleen cells of one immunized mouse were fused with mouse myeloma cells in polyethylene glycol and the positive clones were subcloned 3 times. Results: Two hybridoma cell lines which secrete anti-thyrotropin monoclonal antibodies with high affinity and specificity have been collected. The antibodies were of the IgG1 subclass and their maximum binding with thyrotropin was 60% and 45. 1% respectively. Using competitive binding assay,the antibodies were found to direct against different epitopes of human thyrotropin. Conclusion: The extract of human pituitaries could be used to produce monoclonal anti-pituitary hormone antibodies. The two anti-thyrotropin monoclonal antibodies produced in this study could be used in the establishment of a sensitive measurement of human thyrotropin.展开更多
文摘Hybridoma cells display an increase in antibody productivity following exposure to hypertonic conditions. However, the underlying mechanism is not well understood. In the present study, we hypothesize that the nuclear factor of activated T cells 5 （NFAT5）/tonicity enhancer binding protein （TonEBP） functions to increase the antibody productivity of hybridoma cells. NFAT5 is an osmosensitive mammalian transcription factor. However, its ubiquitous expression in various organs that are not bathed in hypertonic milieu suggests that NFAT5 may also regulate cell growth and function under isotonic conditions. In this study, we examined the expression of NFAT5 in hybridoma cells by Western blot analysis, and found that it increased significantly in hypertonic medium. To further define the function of NFAT5 in hybridoma cells, RNA interference technique was used to downregulate the expression of NFAT5 in SGB-8 cells （a hybridoma cell line）. In isotonic medium, antibody productivity ofhybridoma cells was reduced by downregulation of NFAT5 while cell proliferation was not influenced. The results presented here demonstrate that NFAT5 not only plays an important role in osmotic stress response pathway in hybridoma cells but also is essential for optimal antibody productivity.
基金Partially supported by grants from the National Natural Science Foundation of China(No.3880715)General Logistics Department of PLAand the President Foundation of the First Military Medical University
文摘It has been developed in this laboratory that a serum-free medium, designated asDMI,is adaptive for long-term culture,freezing and resurgence in liquid nitrogen,and cloningof hybridoma and parental myelomas as well as for cell fusions.In this report,it is describedthat the myeloma cells grown in DMI for more than 3 months maintained their biological charac-teristics such as induction of aseites and subcutaneous somatic tumor in BALB/c mice and toler-ance toward 8-azaguanine,ete..However,the ability of the tumor cells to attach to glass wallwas decreased.The selecting assay of these cells in HAT medium(medium containing hypoxan-thine,aminopterin and thymidine)showed that the death time in DMI was similar to that in theconventional 15％ newborn calf serum-supplemented medium(RPS15).The hybridoma cellsadapted in DMI secrete monoclonal antibodies in quantities comparable to those produced inRPS15.
文摘To get the hybridoma cell lines secreting anti-thyrotropin monoclonal antibodies with high affinity and specificity. Methods: BALB/c mice were immunized with extract of human pituitaries. The spleen cells of one immunized mouse were fused with mouse myeloma cells in polyethylene glycol and the positive clones were subcloned 3 times. Results: Two hybridoma cell lines which secrete anti-thyrotropin monoclonal antibodies with high affinity and specificity have been collected. The antibodies were of the IgG1 subclass and their maximum binding with thyrotropin was 60% and 45. 1% respectively. Using competitive binding assay,the antibodies were found to direct against different epitopes of human thyrotropin. Conclusion: The extract of human pituitaries could be used to produce monoclonal anti-pituitary hormone antibodies. The two anti-thyrotropin monoclonal antibodies produced in this study could be used in the establishment of a sensitive measurement of human thyrotropin.
