Construction of Human Recombinant ScFv Phage Libraries from the Advanced Stages of Breast Carcinoma Patients

Advances in the field of antibody engineering, and the emergence of powerful screening technology such as filamentous phage display allowed to generate fully human antibodies with high affinities against virtually any desired target from immune or even naIve human repertoires. As a result, the immunogenicity problems related to applications of nonhuman based recombinant antibodies as therapeutic reagents in human were bypassed. In this study, we constructed large human immunoglobulin libraries from the lymph nodes of breast carcinomas patients in two different formats of single-chain fragments of variable domains (scFv) of antibodies. The heterogeneity of the libraries were tested by restriction enzyme analysis and sequencing on DNA samples of randomly selected colonies. Functional expression of the selected scFv molecules in E. coli was demonstrated by Western blotting. Phage rescue and panning of these libraries against the candidate tumor antigens will lead to the identification of novel human scFvs for tumor detection and pave the way towards the generation of a fully human IgG with desired effector functions for possible future tumor therapy.