In:
The Journal of Immunology, The American Association of Immunologists, Vol. 161, No. 10 ( 1998-11-15), p. 5421-5429
Abstract:
Immunoassays are widely used to determine steroid concentrations. However, they are limited by the specificity of anti-steroid mAbs. We used the phage display system combined with molecular modeling and site-specific randomization to improve the affinity and the fine specificity of an anti-cortisol mAb. Using parsimonious mutagenesis, we have generated a library of mutant Ab fragments (scFv) derived from this Ab by randomizing five amino acids chosen by molecular modeling and Ab-hapten contact structural analysis. Anti-cortisol Ab fragments were selected from the library in the presence of steroid analogues to block cross-reacting binders. Specific elution with free cortisol allowed the recovery of clones with up to eightfold better affinity and fivefold less cross-reactivity than the wild-type scFv. This approach can be applied to any anti-hapten Ab and represents a useful approach for obtaining highly specific Abs for use in steroid immunoassays.
Type of Medium:
Online Resource
ISSN:
0022-1767
,
1550-6606
DOI:
10.4049/jimmunol.161.10.5421
Language:
English
Publisher:
The American Association of Immunologists
Publication Date:
1998
detail.hit.zdb_id:
1475085-5
