Functional Analysis of the Thymic Microenvironment Using Recombinant Single Chain Antibodies

Functional analysis of the thymic microenvironment using recombinant single chain antibodies D.B. Palmer, A. Solomou and M.A. Ritter Department of Immunology, Division of Medicine, Imperial College School of Medicine, Hammersmith Hospital, London, W12 0NN, UK 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.