Preparation of 125I-Labeled Peptides and Proteins with High Specific Activity Using IODO-GEN
The reagent IODO-GEN (1,3,4, 6-tetrachloro-3α,6α-diphenylglycoluril) was first introduced by Fraker and Speck in 1978 (1) and rapidly found widespread use for the radioiodination of both peptides and proteins (2). Although the usual application of the reagent in the laboratory is for introduction of an atom of 125I, IODO-GEN has been used successfully for the preparation of proteins, particularly monoclonal antibodies, labeled with 131I and 123I for use in immunoscintigraphy and positron emission tomography (3-5). This chapter addresses only the use of IODO-GEN to radiolabel pure peptides/proteins in aqueous solution, but iodination of proteins on the cell surface of a wide range of intact eukaryotic and prokaryotic cells (6-7) and on subcellular organelles (8) has been accomplished using the reagent. The advantages of IODO-GEN over alternative reagents previously used for radiolabeling such as chloramine T, lactoperoxidase/ hydrogen peroxide, and 125I-labeled Bolton-Hunter reagent (N-succinimidyl-3 [4-hydroxyphenyl] propionate) are that the reaction is rapid, technically simple to perform, and gives reproducibly high levels of incorporation of radioactivity with minimal oxidative damage to the protein.
KeywordsHPLC Acetonitrile Propionate Polystyrene Tryptophan
- 2.Salacinski, P. R. P., McLean, C., Sykes, J. E. C., Clement-Jones, V. V., and Lowry, P. J. (1981) Iodination of proteins, glycoproteins and peptides using a solid-phase oxidizing agent, 1,3,4,6-tetrachloro-3α,6α-diphenyl glycoluril (Iodogen). Analyt. Biochem. 117, 136–146.PubMedCrossRefGoogle Scholar
- 13.Conlon, J. M. (1991) Measurement of neurokinin B by radioimmunoassay, in Methods in Neurosciences, Vol. 6 (Conn, P. M., ed.), Academic Press, San Diego, CA, pp. 221–231.Google Scholar
- 14.Conlon, J. M. (1991) Regionally-specific antisera to human β-preprotachykinin, in Methods in Neurosciences, Vol. 6 (Conn, P. M.,ed.), Academic Press, San Diego, CA, pp.221–231.Google Scholar