Phage display selection and analysis of Ab-binding epitopes.
The identification and characterization of B cell epitopes by combinatorial phage display peptide analyses is based on the principle that unique peptides can be affinity-purified from an enormous collection of random peptides. Moreover, once selected, the peptide sequence can be elucidated; filamentous bacteriophages have been genetically engineered to incorporate the DNA template corresponding to the peptide displayed on its surface. This unit begins with a discussion of some of the factors that distinguish available libraries. Protocols are then provided for affinity selection of antibody-specific phages, determination of phage titer, confirmation and amplification of positive phages, phage characterization, and construction of custom-tailored phages. The selection protocol in this unit is specific and designed for libraries that are used in the authors' laboratory and are based on the fth1 or fd-tet derived vectors. However, information is included for adapting these protocols to the specific requirements of other phage display libraries.
Enshell-Seijffers
David
D
Tel Aviv University, Israel.
Gershoni
Jonathan M
JM
Tel Aviv University, Israel.
D000906:Antibodies / Q000276:immunology*; D001435:Bacteriophages; D002725:Chloroform / Q000737:chemistry; D004247:DNA / Q000737:chemistry; D018985:Epitopes, B-Lymphocyte / Q000276:immunology*; D004926:Escherichia coli / Q000821:virology; D019151:Peptide Library; D010455:Peptides / Q000276:immunology*; D019800:Phenol / Q000737:chemistry; D019809:Streptavidin / Q000737:chemistry
D000906:Antibodies; D018985:Epitopes, B-Lymphocyte; D019151:Peptide Library; D010455:Peptides; D019800:Phenol; D002725:Chloroform; D004247:DNA; D019809:Streptavidin