Production of human--human hybridomas secreting antibody to sheep erythrocytes after in vitro immunization of peripheral blood lymphocytes.
This report describes the formation of human hybridomas after in vitro immunization of peripheral blood lymphocytes (PBL) with an antigen and fusion of the stimulated lymphocytes with a HAT-sensitive human myeloma cell line, RPMI 8226. PBL were stimulated in vitro with sheep erythrocytes (SRBC) plus fresh human serum. PBL of some donors produced anti-SRBC antibody when they were cultured at 2 X 10(6) cells per well in a 24-well plate with the antigen plus fresh human serum for 7 days. Although lymphocytes of some donors were "low-responders" under the above conditions, they responded to SRBC when they were cultured with not only the antigen plus fresh human serum but also with the culture supernatant obtained after phytohemagglutinin (PHA) stimulation of a mixture of PBL from two donors (MLC-PHA sup). The cells sensitized by this procedure were fused with RPMI 8226 cells. Hybrids secreting IgM or IgG anti-SRBC antibodies were obtained. Additionally the ratio of total IgG-producing hybridomas to IgM-producing ones was higher when the MLC-PHA sup was used at the time of the in vitro immunization.
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D003470:Culture Media; D004912:Erythrocytes / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology*; D007114:Immunization; D008214:Lymphocytes / Q000276:immunology*; D012756:Sheep; D013997:Time Factors
D003470:Culture Media