Solid phase enzyme immunoassay of cyclic adenosine 3',5'-monophosphate. Effect of coating strategy upon assay performance in comparison with radioimmunoassay.
We have evaluated two novel enzyme-linked immunosorbent assays (ELISAs) used to quantitate cyclic AMP. In one assay ELISA plates are coated with antigen consisting of a cyclic AMP-polylysine conjugate. Cyclic AMP samples added to plates are quantified by their ability to decrease the binding and anti-cyclic AMP antibodies to the coated antigen. A second ELISA utilizes a plated anti-immunoglobin technique in which plates are coated first with anti-goat IgG and then with goat anti-cyclic AMP antiserum. Cyclic AMP samples are quantified by their ability to compete with cyclic AMP-peroxidase conjugates for binding to the plated anti-cyclic AMP antibodies. The plated anti-immunoglobin ELISA proved to be somewhat more sensitive than the plated antigen ELISA and was comparable in sensitivity to an automated RIA for measuring cyclic AMP in standards and urine samples. Our data fit with the generalization that optimal ELISA sensitivity is obtained through the use of plated anti-immunoglobins rather than plated antigens. Further they demonstrate the practicality of utilizing small ligand-enzyme conjugates for ELISAs.
Linden
J
J
University of Virginia, Diabetes and Endocrine Research Center, Charlottesville 22908.
Vandenhoff
G E
GE
Taylor
D
D
Finkelstein
G L
GL
D000242:Cyclic AMP / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D066298:In Vitro Techniques; D011863:Radioimmunoassay / Q000379:methods
D000242:Cyclic AMP