Production and assay of antibodies to an activator of adenylate cyclase, forskolin.
Forskolin is a unique diterpene activator of adenylate cyclase which has been extensively used in the study of cAMP generating systems. This report describes the production of antibodies to forskolin and the optimization of two sensitive assay methods for such antibodies. 7-0-Hemisuccinyl 7-deacetyl forskolin, coupled to either human serum albumin or goat IgG, was injected into goats to elicit antibodies to the forskolin hapten. Two assay methods, a radioimmunoassay with [12-3H]forskolin as a tracer and a colorimetric enzyme-linked immunosorbent assay (ELISA) with horse radish peroxidase-labelled rabbit anti-goat IgG as an indicator, were optimized to test for the presence of forskolin antibodies in antisera and isolated IgG fractions. The titers for forskolin antisera were 4000-10000. Both assay methods can be adapted to quantify forskolin and its protein conjugates. The availability of antibodies to this diterpene will be useful in accelerating the understanding of the mechanism of adenylate cyclase activation by forskolin.
Department of Biochemistry, University of Miami School of Medicine, Florida.
L T
LT
Ho
R J
RJ
Ho
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D000936:Antigen-Antibody Complex; D005576:Colforsin / Q000276:immunology* / Q000494:pharmacology; D004789:Enzyme Activation; D004797:Enzyme-Linked Immunosorbent Assay; D006041:Goats / Q000276:immunology; D007074:Immunoglobulin G / Q000302:isolation & purification; D011863:Radioimmunoassay
D000906:Antibodies; D000936:Antigen-Antibody Complex; D007074:Immunoglobulin G; D005576:Colforsin; D000262:Adenylyl Cyclases