Antibody-ribosome-mRNA (ARM) complexes as efficient selection particles for in vitro display and evolution of antibody combining sites.

AUTOR(ES)

He, M

RESUMO

We describe a rapid, eukaryotic, in vitro method for selection and evolution of antibody combining sites using antibody-ribosome-mRNA (ARM) complexes as selection particles. ARMs carrying single-chain (VH/K) binding fragments specific for progesterone were selected using antigen-coupled magnetic beads; selection simultaneously captured the genetic information as mRNA, making it possible to generate and amplify cDNA by single-step RT-PCR on the ribosome-bound mRNA for further manipulation. Using mutant libraries, antigen-binding ARMs were enriched by a factor of 10(4)-10(5)-fold in a single cycle, with further enrichment in repeated cycles. While demonstrated here for antibodies, the method has the potential to be applied equally for selection of receptors or peptides from libraries.

Documentos Relacionados

• In vitro selection and evolution of functional proteins by using ribosome display
• kappa Chain joining segments and structural diversity of antibody combining sites.
• mRNA display: Diversity matters during in vitro selection
• In vitro selection of Jun-associated proteins using mRNA display
• Secondary structure prediction and in vitro accessibility of mRNA as tools in the selection of target sites for ribozymes