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 N(131)IP-containing compounds, as judged from precipitation of radioactivity by salting-out of immunoglobulins. Homogeneous binding is obtained with N(131)IP—polylysine, N(131)IP—ε-amino-n-caproic acid (N(131)IP—aminocap), and other structurally related haptens; non-homogeneous binding is obtained with N(131)IP—bovine serum albumin. Binding to salt-precipitated immunoglobulin of N(131)IP—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 N(131)IP—aminocap. The inhibitions indicate that the iodine contained in NIP, but not the carrier protein, contributes significantly to the binding site.