Radiolabeling of immunoglobulin without loss of antibody activity--use of 14C-phenylisothiocyanate with human cytotoxic antibody.
Radioiodination of immunoglobulins, particularly human cytotoxic IgG, induced a marked loss of antibody activity. Phenylisothiocyanate (PITC) readily reacts with alpha-amino groups and the epsilon-amino groups of lysines to form phenylthiocarbamyl derivatives. Since PITC is commercially available with 14C, 3H, and 35S substitutions, it provided an alternative means for labeling immunoglobulin which preserved antibody activity. There were approximately 80 PITC binding sites per IgG molecule, and 14C-PITC was bound with an efficiency in excess of 80%. With as many as 40 PITC molecules bound per IgG molecule, full cytotoxic activity was retained by both anti-HLA isoantisera and human anti-melanoma autoantisera. Even with 70 to 80 molecules of PITC bound per IgG molecule, over 80% of the antibody activity remained. Radiolabeling of antibody with 3H, 14C, or 35S-substituted PITC will greatly facilitate studies of antibody binding, particularly to cell surface antigens.
D000906:Antibodies / Q000032:analysis*; D001666:Binding Sites, Antibody; D002250:Carbon Radioisotopes; D003601:Cytotoxicity Tests, Immunologic; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism*; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D007553:Isotope Labeling; D011234:Precipitins / Q000032:analysis; D013861:Thiocyanates / Q000378:metabolism*
D000906:Antibodies; D002250:Carbon Radioisotopes; D007074:Immunoglobulin G; D007457:Iodine Radioisotopes; D011234:Precipitins; D013861:Thiocyanates; D004121:Dimethyl Sulfoxide