Time-resolved fluorescence imaging in islet cell autoantibody quantitation

Abstract The prodromal period of insulin-dependent diabetes mellitus (IDDM) is characterized by circulating islet cell autoantibodies (ICA) and other beta cell specific autoantibodies. Despite biochemical characterization of the major beta cell autoantigens insulin, glutamic acid decarboxylase and protein tyrosine phosphatase and development of the respective antibody assays, ICA has remained the standard in IDDM prediction. Conventional ICA quantitation using classic fluorochromes is prone to errors since fluorescence intensity is estimated subjectively using the human eye, which is also unable to differentiate specific signals from non-specific signals and autofluorescence. Using Eu 3+ -chelate labelled anti-human polyclonal IgG (decay time 1000 μ s) as the secondary antibody in time-resolved fluorescence imaging (TRFI), the chelate and autofluorescence signals (typical decay time 65 and IA-2 autoantibody assays combined. For later comparisons, the stained slides may be stored in the light for years without any decrease in specific fluorescence.