Hybridoma cells, human

Vitamins and lipids are often required for animal cells to grow in serum-free medium. Phosphoethanolamine and ethanolamine are key additives that facilitate the growth of the mammary tumor cell line 64024 (Kano-Sueoka and Errick, 1981). In addition, ethanolamine promotes the growth of human lymphocytes and mouse hybridoma cells. Short-term cultures of human diploid lung and foreskin fibroblasts grow in medium that includes among its supplements soybean lecithin, cholesterol, sphingomyelin, and vitamin E. 

Fig. 15.7 Glycosylation of an antibody produced in tobacco plants expressing a human 3(l,4)-galactosyltransferase. As illustrated for Guy sl3 in Fig. 15.4, when the monoclonal antibody Mgr48 is produced in wild type tobacco plants (left panel), its glycosylation is structurally different and more heterogeneous than that of its mammalian counterpart (lower panel). When this antibody is produced in tobacco plants expressing the human galactosyltransferase (right panel), 30% of its N-glycans show terminal N-acetyllactosamine sequences identical to those carried by this antibody when it is produced in hybridoma cells.

Bhunia, A. K., Steele, P., Westbrook, D., Bly, L., Maloney, T., and Johnson, M. (1994). A six-hour in vitro virulence assay for Listeria monocytogenes using myeloma and hybridoma cells from murine and human sources. Microb. Pathog. 16, 99-110. 

As noted, one of the remaining challenges in obtaining human hybridoma cells for generahng human monoclonal antibodies is the lack of suitable human myeloma cell lines to generate stable hybrid cells that can be cloned and expanded indefinitely in culture. Many human hybridoma... 

Koropatnick, 1., 1. Pearson, and IF. Harris, Extensive loss of human DNA accompanies loss of antibody production in heteromyeloma hybridoma cells. Mol Biol Med, 1988.5(2) 69-83. 

The mammalian cell culture technique can be employed to produce clinically important biochemicals such as human growth hormones, interferon, plasminogen activator, viral vaccines, and monoclonal antibodies. Traditionally, these biochemicals had been produced using living animals or extracted from human cadavers. As examples, monoclonal antibodies can be produced by cultivating hybridoma cells in the peritoneal cavity of mice, and the human growth hormone to cure dwarfism can be extracted from human cadavers. However, the quantity obtained from these methods is quite limited for the wide clinical usages of the products. 

Other species of hybridomas, including human, have been produced but are generally created by the use of viruses conferring cellular immortality. Artificial immunization of the donor is often not practical or ethical and so cell lines are often derived from peripheral lymphocytes obtained from individuals naturally immune to the target substance. Some human monoclonal antibody secreting cell lines have been derived from spontaneously occurring myelomas, but this line of approach frequently is unrewarding as the probability that the antibody will be one of interest is remote. 

Both hybridomas and humanized antibody producer cells can be cultivated indefinitely in conventional cultures, usually containing fetal bovine serum in the medium. In the supernatants of these cultures, the quantity of mAbs varies from 20 to 100 pg/ml of protein, depending on the cell and the system of cultivation. 

UMP UTP VO 203 WI-38 uridine monophosphate uridine triphosphate hybridoma cell line Wistar Institute-38 human embryonic lung tissue cell line... 

Many inhibitors of catabolic pathways cause a decrease in cellular heat dissipation. They are therefore valuable tools to indicate the sources of the dissipation and give clues to the relative importance of each pathway in overall metabolic activity (see reviews by Kemp, 1987, 1993 Monti, 1987, 1991). To give a few examples from these reviews, sodium fluoride is a classical inhibitor of glycolysis and it has been shown to substantially reduce heat dissipation by human erythrocytes, lymphocytes, neutrophils, and murine macrophages, indicating the contribution of this pathway to metabolic activity. Cyanide inhibits oxidative phosphorylation by mitochondria at the cytochrome c oxidase complex (site 3) and studies revealed that it decreased heat production in a mouse LS-L929 fibroblast cell line but had no effect on human erythrocytes and neutrophils and murine macrophages, all of which lack mitochondria. Sodium azide inhibits at the same site and so it should come as no surprise that it had no effect on human neutrophils and lymphocytes, but it did reduce heat production by lymphocyte hybridoma cells, which contain... 

Antibodies are expressed by hybridoma cells formed by cell fusion of sensitized animal or human B lymphocytes with myeloma cells, or they are generated by EBV (Epstein-Barr virus) transformation of sensitized B lymphocytes. Other heterologous expression systems such as bacteria, yeast, insect cells, and mammalian cells have also been used for expression of antibodies and their fragments. However, because of renaturation problems, glycosylation, and expression levels, mammalian cells are mostly used for the expression of monoclonal antibodies. More recently, technologies have been extensively developed for the expression of antibodies in transgenic animals and transgenic plants. 

Production of Hybridoma Cell Lines. Eight-week-old female BALB/c mice were injected three times intraperitoneally with 500 xg of human serum albumin (HSA) at 2-week intervals. The first injection was in complete Freund adjuvant (1 1, v/v), and the succeeding two injections were given in incomplete Freund adjuvant (1 1, v/v). The mice were allowed to rest for 30-60 days and injected intravenously with 50 xg of HSA. Four days later, spleen cells from these mice were fused with SP2/0-Agl4 myeloma cells by using polyethylene glycol, as previously described (38). Hy-bridomas of interest were cloned by limiting dilution. 

Von Wedel RJ (1987) Mass culture of mouse and human hybridoma cells in hollow-fibre culture. In Seaver SS (ed.) Commercial Production of Monoclonal Antibodies. A Guide for Scale Up, pp. 159-173. Marcel Dekker, New York. 

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