Immunopharmacology, 7 (1984) %16 Elsevier 9 IMO 00211 Immunochemical Studies on Thymosin: Radioimmunoassay of Thymosin f14 Paul H. Naylor*, John E. McClure**, Bryan L. Spangelo, Teresa L.K. Low and Allan L. Goldstein Department of Biochemistry, The George Washington University School of Medicine, 2300 Eye Street, N.W., Washington, D.C. 20037, U.S.A. (Received 2 June 1983;accepted after revision 15 August 1983) Abstract: Thymosinf14,a peptide with hormonal-like properties first isolated from the thymus gland, can be measured in serum using a newly described radioimmunoassay. The radioimmunoassay utilizes an antibody raised in rabbits against synthetic thymosin t4 conjugated by glutaraldehyde to keyhole limpet hemocyanin. A l z5i_tyrosine.Cl3 analogue of the biologicallyactive C-terminal frag- ment is used as the radioactive tracer. The radioimmunoassayis sensitive in the nanogram range and no cross-reactivitywith common serum proteins is demonstrable. High performance liquid chromatography of serum samples indicates that two thymosin t , cross- reactive species are present in human serum. Levelsin serum range from 450 to 1100ng/ml and decline with age. Key words: Thymosinf14;Thymichormone; Radioimmunoassay Introduction One of the most important advances in medical re- search during the past decade has been the devel- opment of our understanding of the role played by the thymic-dependent immune system in various diseases including the primary immunodeficiency diseases, cancer, infectious diseases, and the au- toimmune diseases (White et al., 1975; Goldstein et al., 1970; Goldstein et al., 1981; Low et al., 1982). Our research program has focused upon studies designed to elucidate the endocrine role of the thy- mus gland and has resulted in the identification of a family of polypeptides from the thymus which have been collectively termed thymosins (Goldstein et al., 1981; Low et al., 1982). Thymosin t4 is a 4963 molecular weight poly- peptide that we have isolated from calf thymus (Low et al., 1981). The amino acid sequence of thy- mosin 1/4 has been determined and published (Low et al., 1982); this polypeptide induces the expression of terminal deoxynucleotidyl transferase in trans- ferase-negative murine lymphocytes in vivo and in vitro (Low et al., 1981). Thymosin 1/4 has also been found to elevate luteinizing hormone releasing hor- mone (LHRH) and luteinizing hormone (LH) in an * To whom reprint requests should be addressed. ** Present address: Section of Allergyand Immunology,Baylor College of Medicine, Houston, TX 77030, U.S.A. Abbreviations: LHRH: luteinizing hormone releasing hormone; LH: luteinizing hormone; RIA: radioimmunoassay; thymosin F5: thymosin fraction five; RIAB: radioimmunoassaybuffer; IgG: immunoglobulinG; cpm: counts per minute; TFA: trifluoroacetic acid; HPLC: high performance liquid chromatography. 0162-3109/84/$03.00 © 1984 Elsevier Science Publishers B.V.