Enrichment and selection of hybrid hybridomas by Percoll density gradient centrifugation and fluorescent-activated cell sorting.

Hybrid hybridomas, producing bi-specific monoclonal antibodies that react with horseradish peroxidase and human IgA1 were isolated by sorting the double-fluorescent cells on single-cell basis after fusion of two hybridomas, previously labelled green or red by octadecylamine-FITC or -TRITC, respectively. The double-fluorescent fused cells were significantly different in AXL (size) and RAS (internal structure) distribution compared with the (non-fused) mono-fluorescent cells. The percentage of double-fluorescent cells and the viability of these cells could be increased by Percoll density gradient centrifugation. As a result, there was an 8-fold increase of total isolated hybrid hybridomas (up to 30% of all tested clones) compared to isolations without Percoll density gradient centrifugation. All the isolated hybrid hybridoma clones had similar amounts of DNA, equal to the sum of the DNA of both parental hybridomas.

[1]  M. Bevan,et al.  Hybrid hybridoma producing a bispecific monoclonal antibody that can focus effector T-cell activity. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[2]  U. Staerz,et al.  Use of anti-receptor antibodies to focus T-cell activity. , 1986, Immunology today.

[3]  D. Scheidegger,et al.  Production of monoclonal antibodies: strategy and tactics. , 1980, Journal of immunological methods.

[4]  R. Colvin,et al.  Bi-specific monoclonal antibodies: selective binding and complement fixation to cells that express two different surface antigens. , 1987, Journal of immunology.

[5]  C. Milstein,et al.  Hybrid hybridomas and the production of bi-specific monoclonal antibodies. , 1984, Immunology today.

[6]  A. Lanzavecchia,et al.  The use of hybrid hybridomas to target human cytotoxic T lymphocytes , 1987, European journal of immunology.

[7]  W. Wright.  The isolation of heterokaryons and hybrids by a selective system using irreversible biochemical inhibitors. , 1978, Experimental cell research.

[8]  M. Gaestel,et al.  Bispecific antibody-producing hybrid hybridomas selected by a fluorescence activated cell sorter. , 1987, Journal of immunological methods.

[9]  S. Person,et al.  A fluorescence enhancement assay of cell fusion. , 1977, Journal of cell science.

[10]  P. Lansdorp,et al.  Human endothelial culture supernatant (HECS): evidence for a growth-promoting factor binding to hybridoma and myeloma cells. , 1981, Journal of immunology.

[11]  M R Suresh,et al.  Advantages of bispecific hybridomas in one-step immunocytochemistry and immunoassays. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[12]  A. C. Cuello,et al.  Hybrid hybridomas and their use in immunohistochemistry , 1983, Nature.

[13]  Philip L. Altman,et al.  Biology Data Book , 1964 .

[14]  P. Lansdorp,et al.  Human endothelial culture supernatant (HECS): a growth factor for hybridomas. , 1980, Journal of immunology.