AbstractA panel of 18 monoclonal antibodies (mAb)Δto insulin have been prepared and used to begin to map antigenic determinants on the insulin molecule. All 18 mAb were of the IgG class, with 14 IgG1, 2 IgG2aand 2 IgG2b. The affinities of these mAb for their immunizing insulin ranged from 1 × 106to 3 × 1081/M. The epitope recognized by three of the mAb, 1, 7 and 16 involves the three residues of the A chain, A 8‐10, the so called A chain‐loop determinant. This A chain loop is one of the most evolutionarily diverse regions of insulins from different species. Another mAb, 10, has been hypothesized to recognize a nearby epitope composed of the A chain residues, A4 and A8 and a B chain residue, B29, that are adjacent on the surface of the insulin molecule. Four of the mAb bind to synthetic B chain. The epitopes recognized by these 4 mAb and the last 10 mAb are unknown but the mAb are grouped according to their ability to bind to different species of insulin or proinsulin. The results of an 18 × 18 matrix analysis of pairs of mAb binding simultaneously to insulin indicate that, despite the finding that some mAb see similar antigenic sites on the insulin molecule, each of the mAb recognizes a unique site on the insulin molecule. Finally, a lower estimate of the number of possible antibodies made to insulin has been calculated to be ≧ 115, a number only 10‐fold lower than the lower limit of antibodies made to dinitrophenyl (DNP) or (4‐hydroxy‐5‐iodo‐3‐nitrophenyl)acetyl (NIP), following hapten
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