The synthesis of a novel radioiodinated ryanodine-O10eq-N-acylamino acylate along with biological data are reported. The affinity of the iodinated product, 7, was comparable to ryanodine, 7.97 nM and 6.47 nM, respectively. Conversion of the non-radioactive iodinated ryanodine analog to the [125I] isotope was accomplished by conversion of 7 to the trimethyltin derivative followed by [125I] exchange using chloramine-T in organic solvent. The radioiodinated ryanodine analog, 9, bound to cardiac membrane preparations in a protein dependent and saturable manner indicating that this analog may represent a useful new tool for the study of ryanodine receptors and that modifications about the C-10 hydroxy group of ryanodine may be carried out without loss in biological activity.