We describe a vitamin B12 radioassay in which oyster toadfish (Opsanus tau) serum is used as the binding protein. The serum is quite stable and has a high capacity and high binding affinity (K greater than 10(12) liter/mol) for vitamin B12. The binding is not significantly affected by temperature, the presence of denatured proteins, or the amount of vitamin B12 present. The radioassay is reproducible (CV 4.4%) within assay, 7.8% between assays) and sensitive (12 ng/liter). Assay accuracy is unaffected by the protein in serum or by reasonable variations in incubation temperature and time.