BACKGROUND

Electrochemical biosensor systems provide an analysis of one or more constituents of blood samples. The biosensor systems include a measurement device having electrical conductor contacts that connect with electrical conductors of the test sensor. The electrical conductors of the test sensor are made from conductive materials, such as solid metals, metal pastes, conductive carbon, conductive carbon pastes, conductive polymers, and the like. The electrical conductors of the test sensor typically connect to working, counter, reference, and/or other electrodes that extend into a sample reservoir. One or more electrical conductors also may extend into the sample reservoir to provide functionality not provided by the electrodes.

A blood sample is introduced into the sample reservoir of the test sensor for analysis. During the analysis, the test sensor includes at least two electrodes in contact with the blood sample referred to as the working electrode and the counter electrode. The sample completes an electrical circuit (allows electrons to flow) between the working and counter electrodes of the test sensor. The test sensor may include additional electrodes or electrical conductors in contact with the blood sample within the sample reservoir that provide a known potential during the analysis (often referred to as a reference or pseudo-reference electrode), sense the presence of the blood sample, and the like. If the counter electrode is active, it may provide a known potential during the analysis.

The blood sample may be drawn from the body of an animal or may be a derivative of such sample, including an extract, a dilution, a filtrate, or a reconstituted precipitate. The analysis performed by the biosensor system of the blood sample determines the content of hematocrit in the blood sample and may determine the presence and/or concentration of one or more analytes, such as alcohol, glucose, uric acid, lactate, cholesterol, bilirubin, ketones, or phenylalanine through the use of an analyte specific ionizing agent.

The measurement device applies an electrical input through the electrical conductor contacts to the electrical conductors of the test sensor. The electrical conductors convey the input through the electrodes into the blood sample present in the sample reservoir. The electrical input may be a potential or current and may be constant, variable, or a combination thereof such as when an AC input is applied with a DC offset. The input may be applied as a single pulse or in multiple pulses, sequences, or cycles.

The oxidation/reduction or redox reaction of one or more constituents of the blood sample generates an electrical output in response to the electrical input. The measurement device measures the electrical output from the test sensor as a current (as generated by amperometry or voltammetry), as a potential (as generated by potentiometry), or as an accumulated charge (as generated by coulometry).

The analyte concentration of the blood sample is determined from an analyte responsive electrical output generated by the redox reaction of the analyte or a species responsive to the concentration of the analyte when the electrical input is applied to the sample. An analyte specific ionizing agent may be added to the sample to enhance electron transfer from the analyte to a second species during the redox reaction. The analyte specific ionizing agent reacts with a single analyte or substantially with a single analyte to provide specificity to a portion of the generated output. An electron transfer mediator may be used to maintain the oxidation state of the analyte specific ionizing agent and more effectively transfer electrons to the working electrode of the test sensor. The measurement device may have the processing capability to measure and correlate the electrical output measured from the test sensor with the presence and/or concentration of one or more analytes in the biological fluid.

While not wishing to be bound by any particular theory, in conventional biosensors for determining blood glucose concentrations for example, glucose may be oxidized by an enzyme, which then transfers the electron to a mediator. This chemically reduced mediator then travels to the working electrode where the chemically reduced mediator is electrochemically oxidized by the potential applied across the working and counter electrode conductors of the test sensor. The oxidized mediator may then return to the reduced enzyme to be re-oxidized to continue the electrochemical reaction of the sample. The amount of mediator being oxidized or reduced by the enzyme and traveling to the working electrode may be correlated to the current flowing between the working and counter electrodes of the test sensor. This current is responsive to the analyte concentration of the sample as the current flowing between the working and counter electrodes is responsive to the reaction of the analyte with the analyte-specific enzyme.

For the quantitative analysis of an analyte in the sample, such as glucose, the electrical output measured at the working electrode should be directly proportional to the diffusion coefficient of the charged analyte or a charged species responsive to the analyte, such as the electron transfer mediator. However, the “hematocrit effect” interferes with this process because the red blood cells in the blood sample (hematocrit or “Hct”) block the diffusion of the charged analyte or the charged species responsive to the analyte to the working electrode of the test sensor. In this way, the hematocrit effect influences the amount of electrical output (e.g. current) measured at the working electrode without any correlation to the amount of analyte (e.g. glucose) in the sample. Thus, for amperometric and voltammetric analysis methods, the current measured by the measurement device at the working electrode of the test sensor varies not only with the analyte concentration of the blood sample, but with the Hct content of the blood sample. As the red blood cells physically interfere with the diffusion of the charged analyte or species responsive to the analyte to the working electrode of the test sensor, the red blood cells may be considered a physical interferent to the analysis of the analyte in the blood sample.

The Hct content of a blood sample varies according to the condition of the animal from which the blood sample is drawn. In humans, the Hct content is normally 39% to 50% by volume (volume of red blood cells/total volume of blood sample*100%) for adult males and 36% to 45% by volume for adult females. Thus, it is desirable to know the Hct content of a blood sample being quantitatively analyzed for an analyte, such as glucose, so that the determined analyte concentration of the blood sample may be corrected based on the Hct content of the blood sample. The Hct content of a blood sample also may be used independently of analyte quantification, such as when blood viscosity or anemia is of interest.

While not wishing to be bound by any particular theory, unlike for an analyte, red blood cells are not believed to undergo a redox reaction during biosensor analysis. Thus, to determine electrochemically the Hct content of a blood sample, a species that can undergo an electrochemical redox reaction at a working electrode is needed. However, to the extent possible, the concentration of this species in the blood sample should not be responsive to that of the analyte concentration of the sample, as different blood samples have different analyte concentrations. Other challenges of Hct analysis include (1) to obtain a Hct responsive output that is large in relation to the non-Hct responsive background output and (2) to obtain a Hct responsive output that provides Hct responsive output amplitudes that are measurably different at different Hct blood sample contents—in combination, the detection sensitivity of the Hct analysis. For example, if the amplitude of the Hct output only changes by 10% or less from 39% to 50% Hct blood sample content, the accuracy of the determined Hct sample content will be poor as there is not enough change in the amplitude of the Hct output to differentiate the different Hct sample contents.

The problem of accurately determining the Hct sample content from small changes in output amplitude is described in U.S. Pat. No. 8,691,072 ('072 patent) where the problem of an only 8% change in output amplitude in relation to a 20% change in Hct sample content is described. Test sensors having this lack of Hct sensitivity issue are described in International Publications 2005/054839 (U.S. Pat. No. 8,088,271) and 2005054840 (U.S. Pat. No. 8,535,497) according to the authors of the '072 patent. The authors of the '072 patent addressed this lack of output amplitude (detection sensitivity) by placing a reductant (ferrocyanide) at the working electrode and an oxidant or reducible species (ferricyanide) at the counter electrode. Thus, different species of a redox pair are present at the working and the counter electrodes of the test sensor. The design of the '072 patent having a reductant at the working electrode and an oxidant at the counter electrode was an improvement over the older '497 patent, where the oxidant ferricyanide was only at or in the vicinity of the counter electrode (so it could be solubilized at the counter electrode)—thus not present at the working electrode. In these systems, a current flow resulted from the test sensor that is inversely proportional to the Hct content of the sample.

In conventional test sensors, electrical conductors transfer electricity from the contacts of the measurement device to the electrodes. Generally, an electrode material or “core” is applied to a portion of the surface of the electrical conductors of the test sensor to form the electrodes of the test sensor by screen printing, sputtering, vapor-deposition, or similar method. The working electrode core material may be palladium, platinum, gold, titanium, carbon, or a mixture of these, for example, which does not undergo oxidation at the potential and duration at which the analysis is performed. A polymer may be added to the electrode core material or used as a film on the core material to seal the electrode core material from the sample. In some instances, the electrode core includes layers of different core materials. The counter electrode is generally formed similarly to the working electrode and may have a single or multiple layers of different core material on the surface of the electrical conductor. The redox species is then applied only to the counter electrode as in the '497 patent or to both the counter and working electrodes as in the '072 patent. Electrodes of this type are inactive.

The 2011/0139634 U.S. Pat. Pub. ('634 publication) to Chou takes a different approach. The '634 publication uses a dedicated reaction zone and a pair of inactive electrodes (silver conductors rendered inactive by a carbon cover to form the electrodes in the reaction zone) to directly measure the Hct content of the sample with an AC electrical input having a DC offset. Thus, a redox species that participates in the electrochemical analysis of the sample is not present. As a DC input will not provide a useful Hct output with inactive electrodes, an AC input is used for Hct determination and the positive correlation between the Hct content of the sample and impedance may be used to determine the Hct content of the sample.

As can be seen from the above description, there is an ongoing need for simple and efficient materials and methods for determining the Hct content of blood samples through electrochemical analysis. The test sensors, electrodes, and methods of the present invention overcome at least one of the disadvantages associated with conventional test sensors and methods.

SUMMARY

In one aspect, the invention provides a test sensor for determining a corrected analyte concentration of a sample including a substrate; at least two electrical conductor contacts and at least two electrical conductors, each electrical conductor contact in electrical communication with an electrical conductor, where the electrical conductor contacts are on the substrate; a lid on the substrate forming a sample reservoir with the substrate, the sample reservoir comprising an inlet port; and at least two pair of electrodes in the sample reservoir in electrical communication with the electrical conductors, where a first electrode pair in the sample reservoir includes an active counter electrode, and a second electrode pair in the sample reservoir includes an analyte specific ionizing agent on a working electrode of the second electrode pair.

In another aspect of the invention, there is a test sensor for determining a physical characteristic of a sample, including a substrate; two electrical conductor contacts and two electrical conductors, each electrical conductor contact in electrical communication with an electrical conductor, where the electrical conductor contacts are on the substrate; a lid on the substrate forming a sample reservoir with the substrate, the sample reservoir comprising an inlet port; and one pair of electrodes in the sample reservoir and in electrical communication with the electrical conductors, where the one pair of electrodes in the sample reservoir includes an active counter electrode.

In another aspect of the invention, there is a method for determining a corrected analyte concentration of a sample including applying a potential of at least 400 mV to a pair of electrical conductor contacts in electrical communication with a pair of electrical conductors in electrical communication with a first electrode pair; generating an electrical output from a second electrode pair responsive to an analyte concentration of a sample, the sample in electrical communication with the second electrode pair; generating an electrical output from a first electrode pair responsive to a physical characteristic of the sample, the sample in electrical communication with the first electrode pair, the first electrode pair including an active counter electrode; correcting the electrical output from the second electrode pair with the electrical output from the first electrode pair; and determining a corrected analyte concentration of the sample. The correcting my include determining an uncorrected analyte concentration of the sample and a physical characteristic content of the sample with previously prepared calibration curves, the previously prepared calibration curves including a relationship between analyte responsive generated output from the second electrode pair and analyte concentration of the sample, and a relationship between physical characteristic generated output from the first electrode pair and physical characteristic content of the sample; and correcting the uncorrected analyte concentration of the sample with the physical characteristic content of the sample.

In another aspect of the invention, there is a method for determining a physical characteristic content of a sample, including applying a potential from 2,000 mV to 6,000 mV to two electrical conductor contacts and two electrical conductors, each electrical conductor contact in electrical communication with an electrical conductor; generating an electrical output from one electrode pair responsive to a physical characteristic of the sample, the sample in electrical communication with the one electrode pair, the one electrode pair including an active counter electrode; and determining a physical characteristic content of the sample from the electrical output.

In another aspect of the invention, there is a system for determining a corrected analyte concentration of a sample including a measurement device having contacts in electrical communication with electrical conductor contacts of a test sensor, each electrical conductor contact in electrical communication with an electrical conductor, where the electrical conductor contacts are on a substrate of the test sensor, the measurement device includes contacts in electrical communication with circuitry including a processor in electrical communication with an electrical input generator and with a storage medium, the test sensor includes a lid on the substrate forming a sample reservoir with the substrate, the sample reservoir comprising an inlet port, the test sensor includes at least two pair of electrodes in the sample reservoir in electrical communication with the electrical conductors of the test sensor, where a first electrode pair in the sample reservoir includes an active counter electrode, and a second electrode pair in the sample reservoir includes an analyte specific ionizing agent on a working electrode of the second electrode pair; the electrical input generator is configured to apply an electrical input of at least 400 mV to the contacts of the measurement device in response to an instruction from the processor; the test sensor is configured to generate an electrical output from the second electrode pair responsive to an analyte concentration of a sample, the sample in electrical communication with the second electrode pair; the test sensor is configured to generate an electrical output from the first electrode pair responsive to a physical characteristic of the sample, the sample in electrical communication with the first electrode pair, the first electrode pair including an active counter electrode; the processor is configured to correct the electrical output from the second electrode pair with the electrical output from the first electrode pair in response to non-transient instructions stored in the storage medium; and the processor is configured to determine a corrected analyte concentration of the sample in response to non-transient instructions stored in the storage medium.

In another aspect of the invention, there is a system for determining a physical characteristic of a sample, including a measurement device having contacts in electrical communication with electrical conductor contacts of a test sensor, each electrical conductor contact in electrical communication with an electrical conductor, where the electrical conductor contacts are on a substrate of the test sensor, the measurement device includes contacts in electrical communication with circuitry including a processor in electrical communication with an electrical input generator and with a storage medium, the test sensor includes a lid on the substrate forming a sample reservoir with the substrate, the sample reservoir comprising an inlet port, the test sensor includes one pair of electrodes in the sample reservoir in electrical communication with the electrical conductors of the test sensor, where the one pair of electrodes in the sample reservoir include an active counter electrode, and the electrical input generator is configured to apply an electrical input from 2,000 mV to 6,000 mV to the contacts of the measurement device in response to an instruction from the processor; the test sensor is configured to generate an electrical output from one electrode pair on the substrate responsive to a physical characteristic of the sample, the sample in electrical communication with the one electrode pair, the one electrode pair including an active counter electrode; the processor is configured to determine a physical characteristic content of the sample in response to non-transient instructions stored in the storage medium.

In another aspect of the invention, there is an active electrode pair for participating in the electrochemical reaction of an analyte in a sample, the active electrode pair including a working electrode, the working electrode consisting essentially of carbon and silver metal, silver metal and silver chloride, and combinations thereof; and a counter electrode, the counter electrode consisting essentially of carbon and silver metal, silver metal and silver chloride, and combinations thereof, where the silver content of the counter electrode is from 70% to 90% (weight/weight).

Other systems, methods, features and advantages of the invention will be, or will become, apparent to one with skill in the art upon examination of the following figures and detailed description. It is intended that all such additional systems, methods, features, and advantages be included within this description, be within the scope of the invention, and be protected by the claims that follow.

