Protein labelling via deoxyribonucleic acid hybridization.
A novel method of radiolabelling antibodies and other proteins is described. A small single-stranded DNA was covalently conjugated to an antibody and labelled by hybridization following the addition of the complementary single-stranded DNA labelled with technetium-99m (99Tc(m)) or indium-111 (111In). Antibody labelling efficiencies were 100% in about 1 h at room temperature with specific activities of up to 30 microCi micrograms-1 of IgG for 99Tc(m). Both diester and thioate DNAs were used. Both the diester- and thioate-labelled antibodies showed complete label stability in 37 degrees C saline. After 24 h in 37 degrees C serum, however, about 40% of the label in the case of the diester antibody was on low molecular weight species--probably labelled catabolites from nuclease degradation of the phosphodiester DNA. In contrast, the 99Tc(m) label on the thioate antibody was immediately and quantitatively bound to serum proteins--probably due to non-specific binding through the sulphur groups. Biodistribution studies in normal mice reflect these in vitro observations: 99Tc(m) on the diester antibody was rapidly cleared through the kidneys, probably as low molecular weight catabolite, while on the thioate antibody, the 99Tc(m) label was predominately deposited in the liver. In conclusion, by modifying with a single-stranded DNA, proteins may be readily labelled with a variety of radionuclides by DNA hybridization. The properties of the radiolabel are strongly influenced by the nature of the DNA.
Hnatowich
D J
DJ
Department of Nuclear Medicine, University of Massachusetts Medical Center, Worcester 01655, USA.
Mardirossian
G
G
Rusckowski
M
M
Winnard
P
P
D000818:Animals; D001483:Base Sequence; D001798:Blood Proteins; D004277:DNA, Single-Stranded / Q000493:pharmacokinetics; D007074:Immunoglobulin G / Q000378:metabolism; D007202:Indicators and Reagents; D007205:Indium Radioisotopes / Q000493:pharmacokinetics*; D007553:Isotope Labeling / Q000379:methods; D051379:Mice; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000493:pharmacokinetics; D013667:Technetium / Q000493:pharmacokinetics*; D013873:Thionucleotides; D014018:Tissue Distribution
D001798:Blood Proteins; D004277:DNA, Single-Stranded; D007074:Immunoglobulin G; D007202:Indicators and Reagents; D007205:Indium Radioisotopes; D009838:Oligodeoxyribonucleotides; D013873:Thionucleotides; D013667:Technetium