title: Increased production of antigen-specific B lymphocytes during in vitro immunization using carrier-specific T helper hybridomas.
abstract: An in vitro method to increase the production of hapten-specific antibody-forming B cells (AFC) using a carrier-specific T helper hybridoma and murine splenocytes is described. Naive splenocytes (6 x 10(6)/ml) are cultured in vitro in the presence of a hapten-carrier conjugate (DNP.OVA) and OVA-specific T helper hybridomas (0.5 x 10(6)/ml). After 4-5 days in vitro immunization (IVI), the maximum number of DNP-specific AFC were found using a spot-ELISA with twice the number of IgM positive cells as IgG positive AFC. The presence of antigen in the form of a hapten-carrier complex and the use of a carrier-specific Th hybridoma resulted in more hapten-specific AFC than when neither antigen nor Th hybridoma were present or when antigen alone or T help alone were used. Also when the hapten was conjugated to a carrier not recognised by the carrier-specific Th hybridoma there were considerably fewer (less than 50%) hapten specific AFC formed. When in vivo primed splenocytes (DNP) were boosted in vitro (IVB) under the same conditions as for IVI most hapten-specific AFC were found on day 4 and both anti-DNP IgM and IgG AFC were increased relative to IVI. Again most AFC were found when hapten was bound to the relevant carrier. In conclusion, carrier-specific T hybridomas can be used in an in vitro immunization procedure with naive or primed splenocytes to increase the frequency of anti-hapten AFC. This method offers an improvement over the current in vitro immunization procedures for the production of monoclonal antibodies.
authors.affiliation: Department of Clinical Immunology, University Hospital, Groningen, Netherlands.
authors.forename: B M
authors.initials: BM
authors.lastname: Schilizzi
authors.forename: B J
authors.initials: BJ
authors.lastname: Kroesen
authors.forename: T H
authors.initials: TH
authors.lastname: The
authors.forename: L
authors.initials: L
authors.lastname: de Leij
mesh_terms: D000818:Animals; D000941:Antigens / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D004136:Dinitrobenzenes / Q000276:immunology; D005260:Female; D006241:Haptens / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology*; D007114:Immunization; D007074:Immunoglobulin G / Q000032:analysis; D007075:Immunoglobulin M / Q000032:analysis; D051379:Mice; D008807:Mice, Inbred BALB C; D010047:Ovalbumin / Q000276:immunology; D006377:T-Lymphocytes, Helper-Inducer / Q000276:immunology*
chemical_list: D000941:Antigens; D004136:Dinitrobenzenes; D006241:Haptens; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D010047:Ovalbumin