Chemical and serological studies with an iodine-containing synthetic immunological determinant 4-hydroxy-3-iodo-5-nitrophenylacetic acid (NIP) and related compounds

Abstract

The synthesis and properties of 4-hydroxy-3-iodo-5-nitrophenylacetic acid (NIP) and several related compounds are described. Conjugates of NIP with proteins are prepared from the azide, synthesized from commercial 4-hydroxy-phenylacetic acid. Sera from rabbits and mice immunized with NIP—ovalbumin or NIP—chicken serum globulin bind N¹³¹IP-containing compounds, as judged from precipitation of radioactivity by salting-out of immunoglobulins. Homogeneous binding is obtained with N¹³¹IP—polylysine, N¹³¹IP—ε-amino-n-caproic acid (N¹³¹IP—aminocap), and other structurally related haptens; non-homogeneous binding is obtained with N¹³¹IP—bovine serum albumin. Binding to salt-precipitated immunoglobulin of N¹³¹IP—aminocap, the hapten of choice for this purpose, provides an assay for antibody measurable at concentrations down to at least M^-9 serum binding capacity (∼0.1 μg antibody/ml). Structurally related compounds and NIP—protein conjugates competitively inhibit binding of N¹³¹IP—aminocap. The inhibitions indicate that the iodine contained in NIP, but not the carrier protein, contributes significantly to the binding site.