Dataset Information

Diabetes Antibody Standardization Program: First evaluation of assays for autoantibodies to IA-2?.

ABSTRACT: Autoantibodies to IA-2? (IA-2?A) are important risk markers of type 1 diabetes. We report the first Diabetes Antibody Standardization Program (DASP) evaluation of IA-2?A assays.Thirteen laboratories from nine countries received coded sera from 50 patients with newly diagnosed type 1 diabetes and 100 healthy blood donors. IA-2?A results were analyzed using receiver operating characteristic (ROC) curves. Concordance of antibody levels was compared using counts per minute (cpm), local and standard curve-derived common units.Median laboratory-assigned sensitivity was 47% (interquartile range [IQR] 45-51), specificity 98% (IQR 95-99), adjusted sensitivity at 95% specificity 50% (IQR 49-53), and area under the ROC curve 0.70 (IQR 0.69-0.73). Use of common IA-2?A units improved concordance between assays compared with local units and cpm (P < 0.0001).IA-2?A assays in multiple laboratories worldwide achieved good concordance and high specificity for type 1 diabetes. IA-2?A are suitable for inclusion in autoantibody testing for risk assessment in prediabetes.

SUBMITTER: Schlosser M

PROVIDER: S-EPMC3198291 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Diabetes Antibody Standardization Program: First evaluation of assays for autoantibodies to IA-2β.

Schlosser Michael M Mueller Patricia W PW Achenbach Peter P Lampasona Vito V Bingley Polly J PJ

Diabetes care 20110916 11

<h4>Objective</h4>Autoantibodies to IA-2β (IA-2βA) are important risk markers of type 1 diabetes. We report the first Diabetes Antibody Standardization Program (DASP) evaluation of IA-2βA assays.<h4>Research design and methods</h4>Thirteen laboratories from nine countries received coded sera from 50 patients with newly diagnosed type 1 diabetes and 100 healthy blood donors. IA-2βA results were analyzed using receiver operating characteristic (ROC) curves. Concordance of antibody levels was compa ...[more]

PMID: 21926293

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Project description:BackgroundCytokine flow cytometry (CFC) or intracellular cytokine staining (ICS) can quantitate antigen-specific T cell responses in settings such as experimental vaccination. Standardization of ICS among laboratories performing vaccine studies would provide a common platform by which to compare the immunogenicity of different vaccine candidates across multiple international organizations conducting clinical trials. As such, a study was carried out among several laboratories involved in HIV clinical trials, to define the inter-lab precision of ICS using various sample types, and using a common protocol for each experiment (see additional files online).ResultsThree sample types (activated, fixed, and frozen whole blood; fresh whole blood; and cryopreserved PBMC) were shipped to various sites, where ICS assays using cytomegalovirus (CMV) pp65 peptide mix or control antigens were performed in parallel in 96-well plates. For one experiment, antigens and antibody cocktails were lyophilised into 96-well plates to simplify and standardize the assay setup. Results ((CD4+)cytokine+ cells and (CD8+)cytokine+ cells) were determined by each site. Raw data were also sent to a central site for batch analysis with a dynamic gating template. Mean inter-laboratory coefficient of variation (C.V.) ranged from 17-44% depending upon the sample type and analysis method. Cryopreserved peripheral blood mononuclear cells (PBMC) yielded lower inter-lab C.V.'s than whole blood. Centralized analysis (using a dynamic gating template) reduced the inter-lab C.V. by 5-20%, depending upon the experiment. The inter-lab C.V. was lowest (18-24%) for samples with a mean of > 0.5% IFNgamma + T cells, and highest (57-82%) for samples with a mean of < 0.1% IFNgamma + cells.ConclusionICS assays can be performed by multiple laboratories using a common protocol with good inter-laboratory precision, which improves as the frequency of responding cells increases. Cryopreserved PBMC may yield slightly more consistent results than shipped whole blood. Analysis, particularly gating, is a significant source of variability, and can be reduced by centralized analysis and/or use of a standardized dynamic gating template. Use of pre-aliquoted lyophilized reagents for stimulation and staining can provide further standardization to these assays.

Dataset Information

Diabetes Antibody Standardization Program: First evaluation of assays for autoantibodies to IA-2?.

Publications

Diabetes Antibody Standardization Program: First evaluation of assays for autoantibodies to IA-2β.

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