Ribosome display: an in vitro method for selection and evolution of antibodies from libraries.
Combinatorial approaches in biology require appropriate screening methods for very large libraries. The library size, however, is almost always limited by the initial transformation steps following its assembly and ligation, as other all screening methods use cells or phages and viruses derived from them. Ribosome display is the first method for screening and selection of functional proteins performed completely in vitro and thus circumventing many drawbacks of in vivo systems. We review here the principle and applications of ribosome display for generating high-affinity antibodies from complex libraries. In ribosome display, the physical link between genotype and phenotype is accomplished by a mRNA-ribosome-protein complex that is used for selection. As this complex is stable for several days under appropriate conditions, very stringent selections can be performed. Ribosome display allows protein evolution through a built-in diversification of the initial library during selection cycles. Thus, the initial library size no longer limits the sequence space sampled. By this method, scFv fragments of antibodies with affinities in the low picomolar range have been obtained. As all steps of ribosome display are carried out entirely in vitro, reaction conditions of individual steps can be tailored to the requirements of the protein species investigated and the objectives of the selection or evolution experiment.
Schaffitzel
C
C
Biochemisches Institut, Universität Zürich, Wintherthurerstr. 190, Zürich, Switzerland.
Hanes
J
J
Jermutus
L
L
Plückthun
A
A
D000818:Animals; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis; D019151:Peptide Library; D012270:Ribosomes
D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D019151:Peptide Library