Effect of different binding proteins on the detection limits and sensitivity of assays based on biotinylated adenosine deaminase.
The properties of binding proteins that control the nature and magnitude of inhibition of the enzyme-ligand conjugates in homogeneous enzyme-linked competitive binding assays were investigated. An assay for biotin that employed adenosine deaminase as the enzyme-label was used as a model system because of the availability of several biotin-specific binders with different characteristics. It was found that the association constant between the ligand and the binding protein, as well as the depth of the binding pocket, affect the response characteristics of the assay. In addition, the reported data suggest that steric-hindrance effects also contribute toward the sensitivity and detection-limit capabilities of the assay.
Barbarakis
M S
MS
Department of Chemistry, University of Kentucky, Lexington 40506-0055.
Daunert
S
S
Bachas
L G
LG
D000243:Adenosine Deaminase; D000818:Animals; D001360:Avidin; D001710:Biotin / Q000032:analysis*; D002352:Carrier Proteins; D002417:Cattle; D005492:Folic Acid; D007202:Indicators and Reagents; D007700:Kinetics; D008832:Microchemistry; D013053:Spectrophotometry / Q000379:methods
D002352:Carrier Proteins; D007202:Indicators and Reagents; D001360:Avidin; D001710:Biotin; D005492:Folic Acid; D000243:Adenosine Deaminase