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Generation of Recombinant Human IgG Monoclonal Antibodies from Immortalized Sorted B Cells
Published on: June 5, 2015
Stimulation of human B cells specific for Candida albicans for monoclonal antibody production
C Davenport1, G F Bottazzo, I Todd
1Department of Immunology, University Hospital, Queen's Medical Centre, Nottingham, UK.
Insights
Researchers developed a new method to create human monoclonal antibodies against Candida albicans. This technique uses specific B lymphocyte stimulation and immortalization for potential infectious disease treatments.
Area of Science:
- Immunology
- Cell Biology
- Infectious Diseases
Background:
- Human monoclonal antibodies are crucial for treating infectious diseases.
- Generating specific human antibodies in vitro presents challenges.
Purpose of the Study:
- To investigate the stimulation and immortalization of human B lymphocytes specific for Candida albicans antigen.
- To establish a method for producing human monoclonal antibodies against infectious agents.
Main Methods:
- An in vitro immunization system using L-leucyl L-leucine methyl ester to remove suppressive cells.
- Culture of remaining cells (CD4+ T cells, B cells, dendritic cells) with antigen and cytokines (IL-2, -4, -6).
- Epstein-Barr virus transformation of activated B cells for antibody-secreting lines.
Main Results:
- An optimal cytokine mixture (IL-2, -4, -6) was identified for antibody production via Elispot assay.
- Established antibody-secreting cell lines specific for anti-Candida antibodies.
- Demonstrated the potential for generating hybridomas or using phage display libraries for antibody production.
Conclusions:
- The developed strategy effectively generates human B lymphocyte lines specific for Candida albicans antigen.
- This method provides a foundation for producing human monoclonal antibodies against infectious agents.
- The overall approach may be broadly applicable for generating human monoclonal antibodies to various pathogens.
Abstract:
The stimulation and immortalisation of human peripheral blood B lymphocytes specific for Candida albicans antigen were investigated. An in vitro immunisation system was employed which involved pretreatment of mononuclear cells with L-leucyl L-leucine methyl ester which removes the suppressive effects of CD8+ T cells, NK cells and monocytes. The remaining cells, CD4+ T cells, B cells and dendritic cells, were cultured with antigen and a mixture of cytokines. A mixture of IL-2, -4 and -6 was found to be optimal for antibody production as determined by an Elispot assay. Transformation of the activated B cells by Epstein Barr virus was found to be optimal after 2 days and lines secreting anti-Candida antibodies were established. These lines could form the basis for specific monoclonal antibody production by generating hybridomas, or by a newly described technique whereby cDNA encoding antibody Fab regions is transferred into phage display libraries. The overall strategy might be generally applicable for the generation of human monoclonal antibodies to infectious agents.
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