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Selected Protocol from "Protein-Protein Interactions", 2nd Edition, E. Golemis, P. Adam.
cAMP Assays
Gouzel Karimova and Daniel Ladant
Unite Postulante de Biochimie des Interactions Macromoleculaires, Departement de Biologie Structurale et Chimie,
CNRS URA 2185, Institut Pasteur, Paris, France
ABSTRACT
cAMP measurements are obtained using an ELISA assay (Harlow and Lane 1988). Commercial radio-immunoassays, or
ELISA kits, to assay cAMP can be purchased from various manufacturers. In our laboratory we use the homemade,
less-expensive ELISA that is described below. In outline, this assay is based on the ability of soluble cAMP in
bacterial extracts to compete with binding of an alkaline phosphatase (AP)-conjugated anti-cAMP antibody to
immobilized cAMP. The surface-bound, immobilized anti-cAMP-AP is inversely correlated to the concentration of
cAMP in the extract. Known concentrations of soluble cAMP are used to calibrate the assay. |
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Products Available for this Protocol
| Protocol Material Description |
Product # |
Product Name |
Add to Cart |
| Buffers, Solutions, and Reagents |
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| BSA |
A3156 |
Albumin, essentially globulin-free from bovine serum |
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| 0.1 M Na2CO3 (pH 9.5) |
S7795 |
Sodium carbonate, SigmaUltra |
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| Dimethyl formamide (DMF) |
D4551 |
N,N-Dimethylformamide |
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| N-Ethoxy-carbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) |
149837 |
2-Ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline |
|
| 50 mM HEPES (pH 7.5) |
H4034 |
HEPES, Biotechnology Performance Certified |
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| NaCl |
S3014 |
Sodium chloride |
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| 0.1% Tween-20 |
P9416 |
TWEEN® 20,for molecular biology, viscous liquid |
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| Cells |
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| Bacterials strains to be assayed for cAMP |
ECACC cell lines |
Cell Lines |
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| Antibodies |
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| Rabbit anti-cAMP antiserum |
A0670 |
Anti-Adenosine 3',5'-Cyclic Monophosphate (cAMP) antibody produced in rabbit, whole antiserum, lyophilized powder |
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