Theoretical limitations of quantification for noncompetitive sandwich immunoassays

Analytical and Bioanalytical Chemistry
Christine F WoolleyTom Taylor

Abstract

Immunoassays exploit the highly selective interaction between antibodies and antigens to provide a vital method for biomolecule detection at low concentrations. Developers and practitioners of immunoassays have long known that non-specific binding often restricts immunoassay limits of quantification (LOQs). Aside from non-specific binding, most efforts by analytical chemists to reduce the LOQ for these techniques have focused on improving the signal amplification methods and minimizing the limitations of the detection system. However, with detection technology now capable of sensing single-fluorescence molecules, this approach is unlikely to lead to dramatic improvements in the future. Here, fundamental interactions based on the law of mass action are analytically connected to signal generation, replacing the four- and five-parameter fittings commercially used to approximate sigmoidal immunoassay curves and allowing quantitative consideration of non-specific binding and statistical limitations in order to understand the ultimate detection capabilities of immunoassays. The restrictions imposed on limits of quantification by instrumental noise, non-specific binding, and counting statistics are discussed based on equilibrium relat...Continue Reading

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Citations

Sep 26, 2017·Nature Nanotechnology·Volker GatterdamChristof Fattinger
Aug 31, 2017·Analytical and Bioanalytical Chemistry·Jie DingMark A Hayes
Mar 11, 2018·Biochemistry. Biokhimii︠a︡·D V SotnikovB B Dzantiev
Mar 4, 2021·Trends in Biochemical Sciences·Luke F Vistain, Savaş Tay
Aug 7, 2021·Journal of Immunoassay & Immunochemistry·Mekbib AstatkeAmy Connolly

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