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Methods. 2013 Mar 15;60(1):27-37. doi: 10.1016/j.ymeth.2012.03.010. Epub 2012 Mar 23.

Cells and cell lysates: a direct approach for engineering antibodies against membrane proteins using yeast surface display.

Author information

1
University of Wisconsin-Madison, Dept. of Chemical and Biological Engineering, 1415 Engineering Dr., Madison, WI 53706, USA.

Abstract

Membrane proteins (MPs) are often desirable targets for antibody engineering. However, the majority of antibody engineering platforms depend implicitly on aqueous solubility of the target antigen which is often problematic for MPs. Recombinant, soluble forms of MPs have been successfully employed as antigen sources for antibody engineering, but heterologous expression and purification of soluble MP fragments remains a challenging and time-consuming process. Here we present a more direct approach to aid in the engineering of antibodies to MPs. By combining yeast surface display technology directly with whole cells or detergent-solubilized whole-cell lysates, antibody libraries can be screened against MP antigens in their near-native conformations. We also describe how the platform can be adapted for antibody characterization and antigen identification. This collection of compatible methods serves as a basis for antibody engineering against MPs and it is predicted that these methods will mature in parallel with developments in membrane protein biochemistry and solubilization technology.

PMID:
22449570
PMCID:
PMC3405166
DOI:
10.1016/j.ymeth.2012.03.010
[Indexed for MEDLINE]
Free PMC Article

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