Selection of antibodies to cell surface determinants on mouse thymic epithelial cells using a phage display library.
The network of thymic epithelium contributes significantly to the thymic stromal cell environment, which plays a vital role in the generation and maturation of thymocytes. Monoclonal antibodies (mAb) have revealed considerable heterogeneity within this epithelial component of the mouse thymic microenvironment, but many of these antibodies recognize epitopes that are located inside the cell and so cannot be used in functional studies. As an alternative approach to isolate antibodies specific to thymic epithelium, we used a phage display library expressing single chain Fv antibodies. For selection, a thymic cell suspension was incubated with the phage display library, and major histocompatibility complex class II positive cells, the majority of which are epithelial, were then specifically selected. Phage bound to these cells were eluted and the selection procedure was repeated for a further five rounds. Immunohistochemical analysis revealed that these phage antibodies show differential staining of thymic epithelial subsets. Flow cytometric analysis of a thymic epithelial cell line using a panel of these antibodies demonstrated that they recognize epitopes on the cell surface. Furthermore, some of these antibodies also labelled human thymic epithelium, suggesting that the epitopes recognized by these antibodies are conserved between human and rodent thymus. Our approach therefore provides a rapid method to select antibodies specific for thymic epithelial cell surface determinants in their native configuration.
Palmer
D B
DB
Department of Immunology, Royal Postgraduate Medical School, Hammersmith Hospital, London, UK.
George
A J
AJ
Ritter
M A
MA
D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D000918:Antibody Specificity; D001435:Bacteriophages / Q000276:immunology*; D002460:Cell Line; D004848:Epithelium / Q000276:immunology; D000939:Epitopes / Q000276:immunology*; D007124:Immunoenzyme Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D008815:Mice, Inbred Strains; D013045:Species Specificity; D013950:Thymus Gland / Q000276:immunology*
D000914:Antibodies, Viral; D000939:Epitopes