Requirements for the generation of memory B cells in vivo and their subsequent activation in vitro for the production of antigen-specific hybridomas.
We have studied the conditions required for the activation in vitro of memory B cells generated in vivo. BALB/c mice were immunised by a single injection of antigen emulsified in Freund's complete adjuvant. Splenocytes were isolated after different time intervals and cultured in a serum-free medium in the presence of antigen and thymocyte-conditioned medium. After 3 days the splenocytes were fused with myeloma cells. A minimum time interval of more than 2 weeks between priming in vivo and stimulation in vitro was required in order to obtain antigen-specific IgG-secreting hybridomas. After a time interval of 4 weeks or longer most of the antigen-specific hybridomas secreted IgGs. During stimulation in vitro the presence of antigen and of T cells was found to be essential for obtaining an antigen-specific IgG response. The addition of thymocyte-conditioned medium enhanced the IgG response.
De Boer
M
M
Laboratory of Biochemistry, University of Amsterdam, The Netherlands.
Ten Voorde
G H
GH
Ossendorp
F A
FA
Van Duijn
G
G
Tager
J M
JM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000145:classification; D001402:B-Lymphocytes / Q000145:classification / Q000276:immunology / Q000378:metabolism*; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D006825:Hybridomas / Q000145:classification / Q000276:immunology / Q000378:metabolism*; D007114:Immunization / Q000379:methods; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000145:classification; D007075:Immunoglobulin M / Q000096:biosynthesis / Q000145:classification; D007156:Immunologic Memory; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D013154:Spleen / Q000166:cytology; D013954:Thyroglobulin / Q000008:administration & dosage / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D013954:Thyroglobulin