Combining phage display and screening of cDNA expression libraries: A new approach for identifying the target antigen of an scFv preselected by phagedisplay

A potential method for identifying new tumor-specific antibody structures a s well as tumor-associated antigens is by selecting scFv phage libraries on tumor cells. This phage display technique involves multiple rounds of phag e binding to target cells, washing to remove non-specific phage and elution to retrieve specific binding phage. Although the binding properties of an isolated tumor-specific scFv can be evaluated by ELISA, FAGS and immunohist ochemistry, it still remains a challenge to define the corresponding antige n. Here, we provide evidence that the target antigen of a given scFv displa yed on phages can be detected in an immobilized lambda phage cDNA expressio n library containing thousands of irrelevant clones. The library contained CD30-negative breast-cancer specific cDNA as well as human CD30 receptor cD NA. The interaction of anti-CD30 scFv phages and their target antigen after blotting onto nitrocellulose filters was documented under defined conditio ns. Screening of different ratios between CD30 receptor and breast cancer s pecific clones (1:1 and 1:200) revealed that the CD30 antigen could be dete cted by anti-CD30 scFv phages using at least 5 x 10(12) plaque forming unit s of filamentous phages per blot These investigations demonstrate that it i s possible to detect the target antigen of a preselected scFv displayed on filamentous phages in lambda phage cDNA expression libraries. (C) 2000 Acad emic Press.