Alternative Display Technologies

Although a number of alternative display technologies have been developed, the two most widely used are yeast display and ribosome display. For yeast display, scFv antibody gene repertoires are fused to the Saccharomyces cell surface protein AgaII.98 Antigen specific scFvs are isolated by subjecting the yeast antibody library to fluorescent activated cell sorting (FACS) using flow cytometry. Yeast display has been used to generate lead antibodies from both immune and non-immune antibody libraries. Its major use, however, has been to increase the affinity of antibodies (antibody affinity maturation). In the most impressive example of affinity maturation using yeast display, the affinity of an scFv was increased more than 1000-fold to 48fM. Potential advantages of yeast compared with phage display include the ability to select more efficiently higher affinity antibodies using flow cytometry and the ability to measure antibody fragment affinity with the antibody in the display format. This obviates the need for antibody fragment expression and purification. Potential disadvantages of yeast display include the lower transformation efficiencies of yeast compared with bacteria.
In ribosome display, scFv antibody gene repertoires encoded by mRNA are translated in vitro in cell-free systems such that the mRNA remaining attached to the ribosome along with the scFv. The ribosome provides the physical link between genotype and phenotype, taking the place of the phage. Antigen binding scFvs are selected on antigen and the mRNA encoding the scFv genes amplified by RT-PCR. Transcription and translation of the scFv DNA provide the display library for the next round of selection. The major advantage of ribosome display is that no cloning is required, resulting in libraries much greater in size than those created by cloning. Ribosome display has been successfully applied to generate scFvs from immune and non-immune libraries and for affinity maturation of existing antibodies.