Comparison of poly- and monoclonal antibodies as labels in a two-site immunochemiluminometric assay for intact parathyroid hormone.
We have evaluated the potential of monoclonal antibodies in the development of a non-isotopic immunometric assay for intact human parathyroid hormone (PTH). The assay has been designed to utilise a chemiluminescent acridinium ester labelled anti-aminoterminal (anti-N) antibody and a solid-phase anti-carboxyterminal antibody in order to measure specifically the intact hormone. In this system the characteristics of the labelled antibody proved crucial to the performance of the assay. A low affinity monoclonal reagent yielded insufficient analytical sensitivity, while a higher affinity monoclonal reagent cross-reacted poorly with the intact molecule relative to the amino terminal PTH fragment to which it was raised. Neither antibody could therefore match the performance of an affinity-purified polyclonal anti-N PTH reagent. These results highlight the problems to be addressed in the selection of suitable reagents for immunometric assay development when specificity and sensitivity are crucial requirements.
Brown
R C
RC
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff.
Aston
J P
JP
St John
A
A
Woodhead
J S
JS
D000818:Animals; D000911:Antibodies, Monoclonal; D000915:Antibody Affinity; D001667:Binding, Competitive; D002417:Cattle; D003429:Cross Reactions; D007118:Immunoassay / Q000379:methods*; D008163:Luminescent Measurements; D010281:Parathyroid Hormone / Q000032:analysis* / Q000276:immunology; D010446:Peptide Fragments / Q000032:analysis / Q000276:immunology; D019379:Teriparatide
D000911:Antibodies, Monoclonal; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D019379:Teriparatide