RADIOIMMUNOASSAY FOR THE CALCIUM-RELEASE CHANNEL AGONIST RYANODINE

A novel photo-activatable derivative of ryanodine, 9-hydroxy-21-(4-azi dobenzoyloxy)-9-epiryanodine, has been synthesized and conjugated to k eyhole limpet hemocyanin for the production of antibodies with high af finity and specificity to ryanodine. The anti-ryanodine antibodies rea cted specifically on immunoblots with the azido-ryanodine compound cov alently conjugated to bovine serum albumin. A radioimmunoassay specifi c for ryanodine was developed using the anti-ryanodine antibodies, and a dissociation constant for ryanodine of 1 nM was determined. Half-ma ximal inhibition constants (IC50) for various ryanodine derivatives we re found to range between 3.2 and 200 nM. These IC50 values correlated very well with the IC50 values obtained for the compounds binding to the skeletal muscle membrane receptor. These antibodies should be usef ul for the characterization of the ryanodine binding site on the sarco plasmic reticulum Ca2+ release channel. (C) 1994 Academic Press Inc.