Method and device for electrochemical immunoassay of multiple analytes

Abstract

A method and device for detection and quantification of biologically significant analytes in a liquid sample is described. The method includes contacting a volume of a liquid sample with predetermined amounts of at least a first and second redox reversible species having redox potentials differing by at least 50 millivolts. At least one of the redox reversible species comprises a liquid sample diffusible conjugate of a ligand analog of an analyte in the liquid sample and a redox reversible label. A predetermined amount of at least one specific binding partner for each analyte to be measured is combined with the sample and current flow is measured at first and second anodic and cathodic potentials and correlated with current flows for known concentrations of the respective diffusible redox reversible species. Diagnostic devices and kits, including such devices and the specified specific binding partner(s) and redox reversible species are also described.

Description

[0001]This application claims benefit of provisional application 60 / 087,576 Jun. 1, 1998.FIELD OF THE INVENTION[0002]This invention relates to a method and device for detection and quantification of biologically significant analytes in a liquid sample. More particularly the invention is directed to a biosensor and method of using same for electrochemical immunoassays of multiple analyte species in a single liquid sample.BACKGROUND AND SUMMARY OF THE INVENTION[0003]Therapeutic protocols used today by medical practitioners in treatment of their patient population requires accurate and convenient methods of monitoring patient disease states. Much effort has been directed to research and development of methods for measuring the presence and / or concentration of biologically significant substances indicative of a clinical condition or disease state, particularly in body fluids such as blood, urine or saliva. Such methods have been developed to detect the existence or severity of a wide va...

AI Technical Summary

Benefits of technology

[0008]One embodiment of the present invention provides a method for measuring multiple analyte species in the same sample, and optimally on the same electrode structure, thus improving the accuracy of the relative measurements. This invention also provides an electrochemical biosensor with capacity to provide improved accuracy through the use of self-compensation. Analyte concentration can be measured / calculated from electrometric data obtained on the same liquid sample with the same electrode structure (the working electrodes), thereby minimizing perturbations due to variability in sample or electrode structure.

[0009]The various embodiments of this invention utilize the principle of diffusional recycling, where a diffusible redox reversible species is alternately oxidized and reduced at nearby electrodes, thereby generating a measurable current. As alternate oxidation and reduction is required for measurement, only electroactive species which are electrochemically reversible are measured thereby eliminating, or at least reducing, the impact or interference from non-reversible electroactive species in the sample. Redox reversible species having different oxidation potentials can be independently measured in a mixture by selecting and bipotentiostatically controlling the oxidizing and reducing potentials for neighboring electrode pairs so that only the species of interest is oxidized at the anode (the electrode with the more positive potential) and reduced at the cathode (the electrode with the less positive or negative potential). When the working electrodes (the anode / cathode arrays) are dimensioned to allow diffusional recycling of the redox-reversible-species at the selected oxidizing and reducing potentials appropriate for that species, a steady state current at the working electrodes where the measurable oxidative and reductive events are taking place, is quickly established through the sample and the electrode structure. The magnitude of the current is proportional to the concentration of the diffusible redox reversible species in the sample. When two or more redox reversible species are utilized, they are selected to have redox potentials differing by at least 50 millivolts, most preferably at least 200 millivolts, to minimize interference between one species and the other in measurements of the respective steady state currents.

Problems solved by technology

However, the use of electrochemical techniques in detecting and quantifying concentrations of such redox reversible species (correlating with analyte concentrations) is not without problem.

These methods suffer from disadvantages including lack of sensitivity and lack of specificity, interference by charging and matrix polarization currents (pulse methods) and electrode fouling due to the inability to apply an adequate overpotential.

Moreover, electrochemical measurements are complicated by interference between the multiplicity of electroactive species commonly extant in biological samples.

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