Construction and guided-selection of mouse-human hybrid Fab phage antibody library against γ-seminoprotein

Objective: To construct a hybrid Fab antibody library and to select human Lc fragment against γ-semino-protein (γ-sm). Methods: Human Lc repertorie gene was amplified by RT-PCR from PBMC of prostate cancer patients and was cloned into pComb3X vector containing murine Fd gene to construct the human-mouse hybrid Fab phage library. Purified γ-sm was used as antigen to screen for 3 rounds and the positive clones were analyzed by Western blot, inhibition ELISA and DNA sequencing. Results: A human-mouse Fab antibody library with good diversity was constructed, with a 1. 2 × 107 CFU size and 90% Lc gene recombinant frequency. After 3 rounds of panning, two hybrid Fab clones specifically against γ-sm were selected, and their apparent binding constant was 71.8% that of the parental mouse antibody E4B7. These two positive clones contained the same light chain with high VL homology to human germline gene IGKV4-1 *01. Conclusion: Specific human Lc fragment against γ-sm can be successfully selected through construction of mouse-human hybrid Fab phage antibody library.