Solid versus liquid phase assays in detection of insulin antibodies. Influence of iodination site on labelled insulin binding.
On type 1 newly diagnosed and on insulin treated diabetic patients, anti-insulin autoantibodies (IAA) and antibodies (IA) having the same specificity are respectively induced. Such immune response may be evaluated either by radiobinding assay (RBA) or enzyme-linked immunosorbent assay (ELISA). Both methodologies have been compared at previous International Workshops, which pointed out discrepancies in results. In this work, IAA/IA prevalence was assessed by displacement RBA and ELISA, in normal subjects, type 2 (treated with hypoglycaemic agents), insulin treated and newly diagnosed type 1 diabetic patients. Results showed a lack of RBA-ELISA agreement. An attempt was then made to determine whether such results were, at least in part, attributable to iodination site in Tyr-A14. For this purpose parallel RBA assays were carried out by using radiolabelled insulin at A14 and A19 Tyr residues. Control sera and samples from insulin treated and type 1 newly diagnosed diabetic patients were tested. Our results suggest that labelling position is not involved in artifactual binding of tracers, at least as a systematic phenomenon. In the majority of cases the variability in RBA-ELISA signal ratios are best explained in terms of differences in the basic principles operating in both methods instead of artifacts due to tracer preparation.
Stumpo
R R
RR
Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Department of Immunology, Argentina.
Llera
A S
AS
Cardoso
A I
AI
Poskus
E
E
D000293:Adolescent; D000328:Adult; D000368:Aged; D001323:Autoantibodies / Q000032:analysis / Q000276:immunology; D001666:Binding Sites, Antibody / Q000276:immunology; D002648:Child; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology; D003924:Diabetes Mellitus, Type 2 / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D007328:Insulin / Q000276:immunology*; D007330:Insulin Antibodies / Q000032:analysis / Q000276:immunology*; D007457:Iodine Radioisotopes; D008297:Male; D008875:Middle Aged; D011869:Radioligand Assay / Q000379:methods*; D014443:Tyrosine
D001323:Autoantibodies; D007328:Insulin; D007330:Insulin Antibodies; D007457:Iodine Radioisotopes; D014443:Tyrosine