Isolation and affinity maturation of hapten-specific antibodies.
More and more recombinant antibodies specific for haptens such as drugs of abuse, dyes and pesticides are being isolated from antibody libraries. Thereby isolated antibodies tend to possess lower affinity than their parental, full-size counterparts, and therefore the isolation techniques must be optimized or the antibody genes must be affinity-matured in order to reach high affinities and specificities required for practical applications. Several strategies have been explored to obtain high-affinity recombinant antibodies from antibody libraries: At the selection level, biopanning optimization can be performed through elution with free hapten, analogue pre-incubation and subtractive panning. At the mutagenesis level, techniques such as random mutagenesis, bacterial mutator strains passaging, site-directed mutagenesis, mutational hotspots targeting, parsimonious mutagenesis, antibody shuffling (chain, DNA and staggered extension process) have been used with various degrees of success to affinity mature or modify hapten-specific antibodies. These techniques are reviewed, illustrated and compared.
Sheedy
Claudia
C
Department of Environmental Biology, University of Guelph, Guelph, Ontario, Canada N1G 2W1.
MacKenzie
C Roger
CR
Hall
J Christopher
JC
D000818:Animals; D000906:Antibodies / Q000276:immunology* / Q000302:isolation & purification*; D000915:Antibody Affinity / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology*; D006241:Haptens / Q000276:immunology*; D006801:Humans; D019151:Peptide Library; D015202:Protein Engineering
D000906:Antibodies; D006241:Haptens; D019151:Peptide Library