Functional Analysis of the Thymic Microenvironment Using Recombinant Single Chain Antibodies
2000
Thymic stromal cells play a crucial role in T lymphocyte development, although the precise molecular interactions that are involved are largely unknown. To investigate these processes, we used a phage display library expressing recombinant single chain antibodies (scFv) [1] and have selected reagents with specificity for molecules expressed on the surface of cortical epithelial cells using either thymic stromal cells [2] or purified microenvironmental molecules as selection substrate. The majority of the scFv recognise evolutionarily conserved epitopes present in the thymus of a wide range of species including human, mouse, rat, pig and chicken. The molecules detected have been analysed by Western blotting; they include the gp200MR6 antigen [3], previously identified only in the human thymus, while others are novel thymic molecules. Some of these scFv influence T cell development in fetal thymic organ culture, indicating that their target molecules play an important role in the interaction between developing thymocytes and the epithelial microenvironment. Others have a direct differentiative effect on epithelium, suggesting that their ligand may play a role in thymocyte-epithelial crosstalk. Our data highlight the power of phage display technology for the selection of specific reagents, for detection of conserved epitopes/molecules on thymic epithelium and for the production of recombinant scFv whose small size enhances tissue penetration and therefore their efficacy in functional analysis.
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