Expression of high-affinity human antibody fragments in bacteria.
Here we describe protocols for the expression of human antibody fragments in Escherichia coli. Antigen-specific clones are identified by soluble fragment ELISA and concentrated by periplasmic preparation. They are then further purified by affinity chromatography. This article provides an overview of expression and purification strategies for human antibody fragments, as well as detailed protocols for the identification of high-affinity binders and for affinity maturation.
Garvan Institute of Medical Research, Darlinghurst, Sydney, New South Wales, Australia.
Romain
R
Rouet
David
D
Lowe
Kip
K
Dudgeon
Brendan
B
Roome
Peter
P
Schofield
David
D
Langley
John
J
Andrews
Peter
P
Whitfeld
Lutz
L
Jermutus
Daniel
D
Christ
D000915:Antibody Affinity; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics
D007128:Immunoglobulin Fragments