In vitro immunization of human B lymphocytes

Informations projet

N° de convention de subvention: BIO2920269

Projet clôturé

Date de début 1 Janvier 1993

Date de fin 30 Juin 1996

Financé au titre de

• Specific research and technological development programme (EEC) in the field of biotechnology, 1990-1994

Coût total € 0,00

Contribution de l’UE € 1 060 000,00

Coordonné par

Danish Cancer Society
Denmark

Résultats exploitables

In vitro immunization of human B-lymphocytes

Technology has been developed which permits the in vitro immunization of human B-lymphocytes. It has been possible to obtain in vitro antigen specific antibody production of the immunoglobulin M (IgM) isotype against primary antigens. These antibodies are usually characteristic of a primary immune response and exhibit affinity constants in the uM range. A method to generate high affinity IgG antibodies is therefore of great interest. Various sources such as tonsils, spleen, peripheral blood and regional lymph nodes have been tested in a number of experiments and it does appear that the most practical source, peripheral blood, can be used for most applications. An aim has been to establish an explant culture system for human lymphoid tissue which could be used for in vitro immunization. The ability of B-cells to present processed antigen on major histocompatibility complex (MHC) class II to specific CD4+ T-cells is of fundamental importance in the induction of antigen specific B-cell responses. The emerging knowledge on the properties of antigenic peptides binding to certain MHC class II allotypes have been considered when constructing so called heterotopes (ie synthetic peptides containing both T and B-cell epitopes). Furthermore, the enhancing effect of certain lipopeptides on antigen presentation, in particular tripalmitoyl cysteine (Pam3Cys) coupled to synthetic peptide antigen have been synthesized. An in vitro cultivation system depending on superantigen (Staphylococcus enterotoxin), which allows activation of single B-cells has been developed. By further manipulating the T-cell compartment, it has been possible to demonstrate an antigen specific isotype switched IgG response in vitro. The in vitro conditions required to obtain the cells secreting high affinity antibodies have also been studied. Initially, the focus has been on the identification of the accessory molecules regulating the growth and differentiation of germinal centre B-cells from human tonsils.