Insulin-like growth factors I and II (IGF-I and IGF-II) were prepared from fetal calf serum and adult bovine serum by gel filtration, immunoaffinity chromatography, chromatofocusing, and reverse-phase high pressure liquid chromatography, and their complete amino acid sequences determined. IGF-I and -II are found in both adult and fetal serum. The sequence of bovine IGF-I is found to be identical to that of human IGF-I, whereas 3 out of 67 amino acid residues are found to be different between bovine and human IGF-II. The differences are located in the C-peptide region of the molecule. Bovine IGF-II shows less than 10% immunological cross-reactivity with antisera against human and rat IGF-II, but is equipotent to human IGF-II in displacing human 125I-labeled IGF-II from human placental receptor. Bovine IGF-I was equipotent to human IGF-I in both radioimmunoassays and radioreceptor assays within the limits of the assay.