Evaluation of islet cell antigen (ICA) 512/IA-2 autoantibody radioassays using overlapping ICA512/IA-2 constructs

Islet cell antigen (ICA)512 also termed IA-2 is a novel autoantigen of type 1 diabetes, which has a tyrosine phosphatase-like domain. We have assessed autoantibody RIAs using a series of ICA512/IA-2 constructs to produce in vitro synthesized S-35-methionine-labeled proteins. Levels of ICA512/IA-2 (256-979, truncated aminoterminus) autoantibodies were strongly correlated with those of the full-length ICA512/IA-2 (1-979) autoantibodies (r = 0.96, P < 0.0001) and ICA512/IA-2 (687-979) autoantibodies (r = 0.98, P < 0.0001). RIAs using these 3 constructs had increased sensitivity relative to our initially reported ICA512 autoantibody RIA (amino acids 389-948, truncated carboxy- and aminoterminus). Only 2 of 38 sera examined in this study reacted with an aminoterminus ICA512/IA-2 (1-577) construct. The mean SD score (so score = (index of unknown sample mean index of controls) using of controls) using the ICA512/IA-2 (256-979) construct was significantly higher than the sn score obtained with other ICA512/IA-2 constructs (P < 0.001). Amongst patients with new-onset diabetes and prediabetic relatives, using RIAs for autoantibodies reacting with ICA512/IA-2 (256-979), insulin, and glutamic acid decarboxylase 65, 98% expressed one or more of these autoantibodies and 78% expressed two or more, whereas no control (n = 208) expressed more than a single autoantibody. A combined ICA512/IA-2 (256-979), glutamic acid decarboxylase 65 autoantibody RIA with differential autoantigen labeling (S-35-methionine, H-3-leucine) has been developed that uses 96-well plate membrane filtration and Top Counter beta counting. Concordance between results of dual and single RIAs was greater than 90%. This simple combined autoantibody assay should facilitate large-scale autoantibody screening.