Rapid Generation of Monoclonal Antibodies from Single B Cells by Ecobody Technology

Abstract

Single B cell sampling following to direct gene amplification and transient expression in animal cells has been recognized as powerful monoclonal antibodies (mAbs) screening strategies. Here we report Ecobody technology which allows mAbs screening from single B cells in two days This technology uses Escherichia coli cell-free protein synthesis (CFPS) for mAb expression. In the CFPS step, we employed our original techniques: (1) ‘Zipbody’ as a modified Fab (fragment of antigen binding) format, in which the active Fab formation is facilitated by adhesive leucine zipper peptides fused at the C-termini of the light and heavy chains; and (2) an N-terminal SKIK peptide tag that can markedly increase protein production. By the Ecobody technology, we demonstrated rapid screening of antigen specific mAbs from immunized rabbits and Epstein-Barr Virus infected human B cells. We further obtained rabbit mAbs in E. coli expression system yielding to 8.5 mg of purified proteins from 1 L bacterial culture.