Hybridoma technology: new developments of practical interest

Samoilovich, Sergio R.; Dugan, Charles B.; Macario, Alberto J. L.

doi:10.1016/0022-1759(87)90147-5

Cited by 33 publications

(11 citation statements)

References 100 publications

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“…A large number of methodological variants for the generation of human MAb secreting hybridomas have been reported (2,3,4,8). However, the frequency of their generation as well as their stability have often been reported as poor (3,5).…”

Section: Discussionmentioning

confidence: 98%

“…We tried to generate improved fusion partners and tried to improve the propagation of hybrids. Generally, fusions with human/mouse heteromyelomas are more stable than with xenogeneic mouse myelomas (4,19). However, only about 1% of the total human chromosomal DNA is incorporated in these heteromyeloma cells (13).…”

Section: Discussionmentioning

confidence: 99%

“…Presently, human MAb can be generated by immortalization of human B lymphocytes either by Epstein-Barr virus transformation and/or fusion to murine, mouse-human heteromyeloma, or lymphoblastoid cell lines (LCL) (2,3,4). However , with respect to efficiency and clone stability, human MAb generation remains far less satisfactory than those of murine MAb (3,5,6,7).…”

Section: Introductionmentioning

confidence: 96%

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Influence of Somatic Cell Hybridization and Human Serum on the Generation and Stability of Human Hybridomas

Lang

Bruderer²

1992

Hybridoma

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We describe an approach that allows the generation of stable hybridomas secreting antigen specific human IgG antibodies with an efficiency comparable to that of the generation of IgM and IgA secreting hybridomas. This was achieved by evaluating means to increase the frequency of human hybridoma formation and the stability of the generated hybridoma cells when subjected to conditions for large scale growth. To this end, we generated new fusion lines with an increased human DNA content and modified the culture system. However, the application of these new fusion lines primarily resulted in unstable giant cells. As a consequence, we evaluated whether the viability of newly formed hybrids between existing fusion lines and lymphoblastoid cell lines might be improved. In an attempt to provide as many components necessary for the growth of antibody secreting hybridomas as possible, we propagated fused cells in medium supplemented with human serum. Our results show that with this approach the frequency of initially growing hybrids was significantly increased. Furthermore, only in culture medium supplemented with human serum was it possible to obtain stable IgG secreting clones.

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Section: Discussionmentioning

confidence: 98%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

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Influence of Somatic Cell Hybridization and Human Serum on the Generation and Stability of Human Hybridomas

Lang

Bruderer²

1992

Hybridoma

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“…With regard to the current technology to fuse antibody-producing B cells with myeloma partner cells, the most popular technique is polyethylene glycol (PEG)-mediated cell fusion. PEG-fusion gives good yields of hybridomas if carefully controlled by following a detailed procedure, but the uncertainty with respect to the outcome of individual experiments still remains because the success of fusion depends, to some extent, on minute details which are difficult to standardize (1,4). Successful improvements of the fusion technique with electrofusion (2, 3, 4) and immunochemical methods (5, 6) have been reported, but none of these techniques have been widely adopted (1).…”

Section: Introductionmentioning

confidence: 98%

“…PEG-fusion gives good yields of hybridomas if carefully controlled by following a detailed procedure, but the uncertainty with respect to the outcome of individual experiments still remains because the success of fusion depends, to some extent, on minute details which are difficult to standardize (1,4). Successful improvements of the fusion technique with electrofusion (2, 3, 4) and immunochemical methods (5, 6) have been reported, but none of these techniques have been widely adopted (1). Recently reported techniques (7,8) to generate MAbs by use of recombinant DNA methodology show promise, but at present these techniques are still at an experimental stage.…”

Section: Introductionmentioning

confidence: 99%

Production of Monoclonal Antibodies by the Use of pH-Dependent Vesicular Stomatitis Virus-Mediated Cell Fusion

Nagata¹,

Yamamoto²,

Ueno³

et al. 1991

Hybridoma

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To explore an alternative fusion method to generate monoclonal antibody (MAb)-producing hybridomas, we have tested whether pH-dependent vesicular stomatitis virus (VSV)-mediated cell fusion is applicable for generation of hybridomas. VSV-mediated cell fusion induced by acid treatment resulted in several specific MAb-producing hybridomas, almost all of which stably produced MAbs. This method is quite simple and offers a potential alternative to other fusion methods. In addition, the fusogenic proteoliposomes prepared by the reconstitution of the solubilized glycoprotein of VSV were also demonstrated to have the ability to generate MAb-producing hybridomas by the same pH-dependent cell fusion.

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Selective cloning of hybridoma cells for enhanced immunoglobulin production using flow cytometric cell sorting and automated laser nephelometry

et al. 1990

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Techniques for selective cloning of murine hybridoma cells by flow cytometric cell sorting and use of automated laser nephelometry to determine the resultant clones' immunoglobulin secretion levels are described. Using a commercially available attachment to a fluorescenceactivated cell sorter, individual hybridoma cells were successfully distributed into microtiter wells in an automated manner based on their forward angle light scatter properties and their reaction to fluorescein-conjugated anti-mouseIgG. The techniques were used to estimate successfully the frequency of immunoglobulin-secreting cells in established cultures. In addition, heterogeneity of cell surface immunoglobulin expression was observed and utilized as a criterion for flow sorting of new hybridoma variants. In these studies, clones derived from high (anti-IgG) intensity sorting regions yielded cultures with enhanced immunoglobulin secretion levels, as determined by automated laser nephelometry. Furthermore, the surface immunoglobulin phenotype of the derived clones was conserved in subsequent progeny. Finally, it was established that inclusion of propidium iodide in the hybridoma cell sorting mixtures improved cloning efficiency by facilitating enhanced discrimination and elimination of nonviable cells. Our results indicate that flow cytometric-assisted single cell depositioin provides positive attributes of severall traditional hybridoma cloning techniques and, in addition, furnishes a tool for steering the cloning process toward selection of enhanced immunoglobulin producing cultures.

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Hybridoma technology: new developments of practical interest

Cited by 33 publications

References 100 publications

Influence of Somatic Cell Hybridization and Human Serum on the Generation and Stability of Human Hybridomas

Influence of Somatic Cell Hybridization and Human Serum on the Generation and Stability of Human Hybridomas

Production of Monoclonal Antibodies by the Use of pH-Dependent Vesicular Stomatitis Virus-Mediated Cell Fusion

Selective cloning of hybridoma cells for enhanced immunoglobulin production using flow cytometric cell sorting and automated laser nephelometry

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