[The in vitro induction of antibody responses of human lymphocytes and its application to hybridoma production].
Sources of immunized lymphocytes constitute one of the main obstacles in the production of human monoclonal antibodies. We tried to get them through in vitro immunization. Cells from excised tonsils or trauma spleens were used for the induction of antibody responses in vitro. Antibodies to different antigens including sheep red blood cells, ovalbumin, tetanus toxoid, and hepatitis B surface antigens were induced in 7-14 days' cultures. Taking tetanus toxoid as antigen, we analysed the various factors required for antibody induction with statistics analysis, which included cell separation method, T cell conditioned medium, antigen dosage, serum content, and concentration of mitogen PWM and LPS. The results showed: (1) The cell separation method influenced the antibody production significantly in comparison with other factors. It signified that immune cells' combination was the most influential factor. (2) Serum also constituted quite important influencing factor especially in the later period of culture. However, it did not make much differences if it attended 10% or so. The antigens and mitogens tended to be used at low concentration. (3) Due to the significant variation among individuals and among different antigens, it is suggested to set up the culture system with some flexibility so as to adapt to the variation in cells and antigens from different sources. The present culture system we use includes nylon wall column separation of cells, suitable range of antigens (three doses instead of one), and either 10% T cell conditioned medium or a mixture of 1 microgram PWM/ ml + 0.1 microgram LPS/ml. The human B lymphocytes stimulated in vitro with tetanus toxoid were used for the construction of human hybridomas.
Yeh
M
M
Shanghai Institute of Cell Biology, Academia Sinica.
Sun
P
P
Ji
Y
Y
Hashizume
S
S
Kamei
M
M
D000818:Animals; D000917:Antibody Formation; D001402:B-Lymphocytes / Q000276:immunology*; D002459:Cell Fusion; D002478:Cells, Cultured; D006801:Humans; D006825:Hybridomas / Q000276:immunology*; D007114:Immunization; D007074:Immunoglobulin G / Q000378:metabolism; D051379:Mice; D013745:Tetanus Toxoid / Q000276:immunology
D007074:Immunoglobulin G; D013745:Tetanus Toxoid