Advantages and pitfalls of radioimmune and enzyme linked immunosorbent assays of insulin antibodies

Sodoyez-Goffaux, F; Koch, Michel; Dozio, Nicoletta; Brandenburg, Dietrich; Sodoyez, J. C.

doi:10.1007/bf00278754

Cited by 58 publications

(45 citation statements)

References 34 publications

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“…By contrast, in the presence of high affinity antibodies, EIA signal is maximum and independent of small differences of affinity. As already discussed, EIA is not appropriate to study species specificity of (high) affinity antibodies circulating in insulin-treated diabetic patients [18]. Direct evidence for differential sensitivity of the two assay systems to avidity of the antigen-antibody reaction was recently obtained using monoclonal insulin antibodies raised in mice (unpublished data).…”

Section: Discussionmentioning

confidence: 99%

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Clonally restricted insulin autoantibodies in a cohort of 2200 healthy blood donors

et al. 1990

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Summary. Serum samples of 2200 blood donors were screened for anti-insulin IgG by enzyme-linked immunosorbent assay. Specificity of detected antibodies was verified by competition with an excess of insulin and observation that saturated anti-insulin IgG were no longer measurable. The upper limit of measured signal in the population was defined as the mean plus three SD. In the direct assay, 32 sera were positive. Among these, 22 (1%) contained saturable insulin antibodies (true positive) and 10 were non-saturable and considered as non-insulin-specific. The positive blood donors were requested to answer a questionnaire and according to their answers, none had ever received insulin, was a first degree relative of a Type i (insulin-dependent) diabetic patient or had experienced a hypoglycaemic episode. None of the 22 true positive sera detected by enzyme-immunosorbent assay bound 12M-insulin to any significant extent. The nine sera yielding the highest signal were further characterized with regard to heavy and light chains as well as species specificity of ligand. Anti-insulin IgG from healthy blood donors contained only one heavy (71 2/9; 73 7/9) and one light (~c 8/9; s 1/9) chain. Three sera were human insulin specific; two were non-species-specific; the other four bound insulin in the order human = porcine > bovine. These results indicate that low affinity clonally restricted antibodies against insulin are present in unselected blood donors with a prevalence of 1%.

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Section: Discussionmentioning

confidence: 99%

“…Detection of insulin antibodies is limited by the sensitivity and the specificity of the assay technique, Furthermore, as recently discussed, currently available assay techniques do not always yield concordant results [18].…”

Section: Introductionmentioning

confidence: 99%

Clonally restricted insulin autoantibodies in a cohort of 2200 healthy blood donors

et al. 1990

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“…Therefore, all sera were also in cubated with excess soluble insulin in the displace ment assay. It has been suggested that ELISA is more efficient than liquid-phase radiobinding assays in determining antibodies with low avidity for the anti gen, a property commonly found in autoantibodies [16][17][18],…”

Section: Discussionmentioning

confidence: 99%

IgG-Class Insulin Autoantibodies in Autoimmune Thyroid Disease

Ng¹,

Thai²,

Lui³

et al. 1990

Int Arch Allergy Immunol

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We studied the incidence of insulin autoantibodies (IAA) in 97 patients with autoimmune thyroid disease with an enzyme-linked immunosorbent assay. The sera were also tested for islet cell antibodies (ICA) and thyrotropin receptor antibodies (TRAb). IAA as assessed by a standard deviation score were present in 7 patients (7.2%). None of the patients were seropositive for ICA. Data from this random sampling and a retrospective study of 12 patients on antithyroid drug treatment indicated that the presence of IAA was not associated with TRAb.

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“…Serum samples from 10 Type I diabetic patients treated with exogenous insulin with IA, and samples from 10 normal healthy controls without IA, detected by either RBA or EIA at different titers, were investigated (30,35,52,53). For comparison with IEM assay, only EIA results of IA measurement are considered in this study.…”

Section: Seramentioning

confidence: 99%

Immunoelectron microscopy: a new method for detection of insulin antibodies.

Saccomanno

Taccagni

Bosi

et al. 1993

J Histochem Cytochem.

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The aim of the present study was to set up a sensitive technical alternative to the classical procedures for detection of human insulin antibodies. We developed a method of post-embedding immunoelectron microscopy (IEM) using as the substrate fresh human pancreas, embedded in acrylic resin to maintain its antigenic structure. The antigen was insulin within the mature secretory granule. Serum samples obtained from 10 patients with insulin antibodies detected at various titers by either radio binding assay (RBA) or enzymatic immunoassay (EIA) were incubated for 2 hr at 37 degrees C at dilutions of 1:25, 1:100, 1:400, 1:1600, 1:6400, 1:25,600, and 1:102,400. The electron microscope photographs were analyzed by computerized morphometry and the number of protein A-gold-IgG complexes was calculated per micron2 of insulin granule. IEM results were compared with those obtained with EIA. The specificity of both techniques towards insulin was assessed as the difference between the signals (number of gold particles per micron2 of insulin granule in IEM or optical density > or = 0.193 in EIA) with and without excess insulin. Sensitivity was defined as the detection limit of the assay. In all the 10 sera investigated, IEM was more sensitive, with a 12- to 40-fold lower detection limit than EIA. IEM, with native insulin granules as substrate, is a specific, reproducible, and sensitive method for detection of human serum insulin antibodies. These findings also suggest IEM as a procedure potentially suitable for identifying antigen specificity of autoantibodies circulating at low concentration.

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Advantages and pitfalls of radioimmune and enzyme linked immunosorbent assays of insulin antibodies

Cited by 58 publications

References 34 publications

Clonally restricted insulin autoantibodies in a cohort of 2200 healthy blood donors

Clonally restricted insulin autoantibodies in a cohort of 2200 healthy blood donors

IgG-Class Insulin Autoantibodies in Autoimmune Thyroid Disease

Immunoelectron microscopy: a new method for detection of insulin antibodies.

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