KR20040007028A - Electrochemical test strip having a plurality of reaction chambers and methods for using the same - Google Patents

Abstract

PURPOSE: A method of using an electrochemical test strip having a plurality of reaction zones bound by opposing electrodes to detect an analyte of a physiological sample is provided. Also provided is a kit for measuring the concentration of the analyte of the physiological sample. The electrochemical test strip can be used in the detection of various analytes in a sample, in particular glucose. CONSTITUTION: The electrochemical test strip(20) includes: 2 to 25 reaction zones bound by opposing electrodes separated by a thin spacer layer; and a reagent composition present in each reaction zone, wherein the reagent composition can be the same or different. Each reaction zone has a separate fluid channel(22), or the reaction zones have separate channels that merge into a single channel to provide a mutual exchange of fluids between reaction zones and an external environment of the test strip. The method of measuring the concentration of an analyte using the electrochemical test strip comprises the steps of: applying a physiological sample to the electrochemical test strip; detecting an electric signal in the reaction zones using the electrodes; and relating the detected electric signal and the amount of the analyte of the sample.

Description

Electrochemical test strip having a plurality of reaction chambers and methods for using the same

FIELD OF THE INVENTION The field of the present invention relates to analyte measurements, in detail electrochemical measurements of analytes, and more particularly electrochemical measurements of blood analytes.

The importance of detecting analytes in physiological body fluids, such as blood or blood-derived products, is of increasing importance in recent conferences. Analyte detection assays find use in a variety of applications, including clinical laboratory tests, in-home tests, and the like, and these test results play a major role in diagnosing and managing various disease states. Such analytes include glucose, cholesterol, and the like for the management of diabetes. As the importance of analyte detection grows in this way, various analytical detection protocols and equipment for clinical and home use have been developed.

One of the methods used for analyte detection is an electrochemical method. In this method, an aqueous solution sample is placed in a reaction zone in an electrochemical cell comprising two electrodes, a reference electrode and a working electrode, wherein the electrode has an impedance that imparts compliance to the ammeter measurement. By reacting the component to be analyzed directly with the electrode or directly or indirectly with the redox reagent, an amount of oxidizing (or reducing) substance corresponding to the concentration of the component to be analyzed, ie, the analyte, can be produced. The amount of oxidative (or reducing) material present is then measured electrochemically and correlated with the amount of analyte present in the initial sample.

Because of the wide applicability of detection protocols by electrochemistry, there is a continuing interest in the formulation of new equipment and methods in this field.

Related literature

The U.S. patents include 5,269,891, 5,830,170, 5,834,224, 5,942,102, and 5,972,199. Other relevant patent documents include WO 99/49307, WO 97/18465 and GB 2 304 628. Other relevant references include: Dalmia et al., J. Electroanalytical Chemistry (1997) 430: 205-214; Nakashima et al., J. Chem. Soc. (1990) 12: 845-847; And Palacin et al., Chem. Mater. (1996) 8: 1316-1325.

Electrochemical test strips and methods for using them in the detection of analytes in physiological samples are provided. The test strip has a plurality of reaction zones defined by opposing metal electrodes separated by a thin spacer layer. The reagent composition present in each reaction zone may be the same or different. In addition, each reaction zone may have a separate fluid entry passage, or two or more reaction zones may have fluid entry passages that combine into a single passage. The electrochemical test strips can be used for the detection of a wide variety of analytes, and are particularly suitable for use in the detection of glucose.

1 shows an electrochemical test strip according to the prior art.

2-4 show an electrochemical test strip according to the invention.

5 shows an electrochemical test strip according to the invention in use.

Electrochemical test strips and methods for using them in the detection of analytes in physiological samples are provided. The test strip has a plurality of reaction zones defined by opposing metal electrodes separated by a thin spacer layer. The reagent composition present in each reaction zone may be the same or different. In addition, each reaction zone may have a separate fluid entry passage, or two or more reaction zones may have fluid entry passages that combine into a single passage. The electrochemical test strips can be used for the detection of a wide variety of analytes, and are particularly suitable for use in the detection of glucose.

Before the present invention is described in more detail, it is to be understood that the invention is not limited to the specific aspects of the invention described below, but modifications may be made to the specific aspects, which also fall within the scope of the appended claims. do. Also, it is to be understood that the terminology used is for the purpose of describing particular embodiments only, and is not intended to be limiting. Instead, the scope of the invention will be established by the appended claims.

In this specification and the appended claims, unless expressly stated to the contrary, the singular encompasses the plural. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

Electrochemical test strip

As summarized above, the electrochemical test strip of the present invention is characterized by having multiple reaction zones. More specifically, the electrochemical test strip consists of two opposing metal electrodes separated by a thin spacer layer, which components define a plurality of separate reaction zones or zones, ie chambers in which the reagent system is installed. Since the electrochemical test strips have multiple reaction zones, they comprise two or more different reaction zones, and in certain embodiments include three to five or more different reaction zones. The number of different reaction zones in the electrochemical test strips according to the invention generally ranges from about 2 to 25, typically about 2 to 15, more typically about 2 to 10, in many embodiments. The different reaction zones of the test strips can generally be arranged in any convenient way on the test strips, in many embodiments they are arranged parallel to the base of one end of the strip, as shown in FIG. 2. . In FIG. 2, the electrochemical test strip 20 has a plurality of reaction zones shown as passages 22. This is in contrast to the conventional electrochemical test strip shown in FIG. 1 with one reaction zone 11.

As summarized above, the test strip consists of a working electrode and a reference electrode separated by a spacer layer arranged to define a plurality of reaction zones or regions of the strip. Each of the elements, namely the working electrode and the reference electrode, the spacer layer, and the reaction region, are described in more detail separately below.

electrode

As noted above, the electrochemical test strip includes a working electrode and a reference electrode. In general, the working electrode and the reference electrode are arranged in the form of an elongate rectangular strip. Typically, the electrode is about 1.9 cm to 5.5 cm in length, typically about 2 to 4.0 cm. The width of the electrode is about 0.20 to 1.0 cm, typically 0.31 to 0.67 cm. The electrode typically has a metal thickness in the range of about 10 to 300 nm, typically about 13 to 60 nm.

The working electrode and the reference electrode, at least the surface of the electrode facing the reaction region of the strip is a metal, such metals are palladium, gold, platinum, silver, iridium, carbon, indium tin oxide doped, stainless steel , Nichrome, and the like. In many embodiments, the metal is gold or palladium. In principle, the entire electrode can be composed of metal, and each electrode can comprise an inert support material on the surface where a thin layer of the metal component of the electrode is present. In these more common aspects, the thickness of the inert support is typically about 51 to 356 μm, typically about 25 to 254 μm, and the thickness of the metal layer is typically about 10 to 150 nm, typically about 15 to 60 nm (e.g., spatter). Sputtered metal layer). Any support that is convenient for use may be used for the electrode, wherein the material is a rigid material capable of providing structural support to the electrode and again to the electrochemical test strip as a whole. Suitable materials that can be used as the support substrate include plastics such as PET, PETG, polyimide, polycarbonate, polystyrene, silicone, ceramics, glass and the like.

The working electrode and reference electrode described above can be fabricated using any protocol that is convenient for use. Representative protocols include, first, a metal electrode preparation that sputters a metal layer of sufficient thickness onto the surface of the inert support.

Spacer layer

The electrochemical test strip is characterized by being separated by only a short distance, with the working electrode and the reference electrode facing each other, wherein the distance between the working electrode and the reference electrode in the reaction zone or region of the electrochemical test strip is It is very small. This minimal spacing between the working electrode and the reference electrode in the test strip is due to the presence of a thin spacer layer sandwiched or positioned between the working electrode and the reference electrode. The thickness of this spacer layer is generally about 12 to 500 μm, typically about 50 to 153 μm.

A feature of the strip is that the spacer layer is cut to provide a plurality of reaction zones or zones as described above, wherein each reaction zone has one or more inlet or entry passages to the reaction zone, and generally There is an outlet or discharge passage from the reaction zone, which provides for the interaction of the fluid between the interior of the reaction zone and the external environment of the strip. The spacer layer of the strip shown in FIGS. 2 to 4 provides a straight passageway, which is an entry and exit passage, acting as a reaction zone, but only other forms, for example an annular reaction zone cut into side inlets and outlets, are possible. In addition, other forms are possible, such as squares, triangles, rectangles, irregularly shaped reaction zones, and the like. The spacer layer may be made of any material that is convenient for use, and representative examples of suitable materials include PET, PETG, polyimide, polycarbonate, and the like, and the surface of the spacer layer may be bonded to their respective electrodes. Can be processed, thereby maintaining the structure of the electrochemical test strip. It is particularly useful to use die-cut double-sided adhesive strips as spacer layers.

Reaction zone

As mentioned above, the electrochemical test strip comprises a plurality of reaction zones or regions defined by the above-mentioned components, the working electrode, the reference electrode and the spacer layer. Specifically, the working and reference electrodes define the top and bottom of the reaction zone and the spacer layer defines the walls of the reaction zone. The volume of the reaction zone is at least about 0.1 μl, typically at least about 0.3 μl, more typically at least about 0.5 μl, wherein the volume may be at least 10 μl. As mentioned above, each reaction zone generally includes one or more inlets (entry passages), and in many embodiments includes outlets (outlet passages). The cross-sectional area of the inlet and outlet may vary within a range sufficient to effectively provide the inlet and outlet of fluid from the reaction zone, but is generally about 9 x 10 ^-5 to 5 x 10 ^-3 cm ² , typically about 1.3 x 10 ^-3 to 2.5 x 10 ^-3 cm ² .

In certain aspects of the present invention, each reaction zone or zone has its own separate entry and exit passageway. As a result, in these aspects the number of different entry passages is equal to the number of different reaction zones of the strip. In another aspect, two or more entry passages may be combined into a single passage before exiting the strip, such that fluid may be introduced from one inlet into two or more different reaction zones. In other words, a single entry passage branches into two or more sub-paths and enters different reaction zones of the strip. The spacer layer of the strip is designed to provide the desired entry passage pattern, for example separate passages or merged passages.

Reagent compositions or systems necessary or desired for detection of analytes using the strip are present in each of the reaction zones. In certain embodiments, for example multipurpose strips, the reagent system is the same in all other reaction zones of the strip. In other aspects, for example when the strip is used to analyze panels or multiple different analytes simultaneously, the reagent composition will be different in individual reaction zones or zones. In other words, two or more different reagent compositions will be present in different reaction zones of the strip, and the number of different reagent compositions may be as many as different reaction zones or zones of the strip.

In many embodiments, the one or more reagent compositions are redox reagent systems, which are provided for the species detected by the electrodes and can therefore be used to infer the concentration of the analyte in the physiological sample. Redox reagent systems present in the reaction zone typically include one or more enzyme (s) and mediators. In many embodiments, the enzyme member (s) of the redox reagent system is one enzyme or multiple enzymes that work together to oxidize the analyte of interest. In other words, the enzyme component of the redox reagent system consists of a single analyte oxidase or a collection of two or more enzymes that work together to oxidize the analyte of interest. Examples of such enzymes include oxidase, dehydrogenase, lipase, kinase, diaphorase, quinoprotein, and the like.

The specific enzyme present in the reaction zone is determined by the specific analyte to be designed by detecting the electrochemical test strip, and representative enzymes include glucose oxidase, glucose dehydrogenase, cholesterol esterase and cholesterol oxidase. , Lipoprotein lipase, glycerol kinase, glycerol-3-phosphate oxidase, lactate oxidase, lactate dehydrogenase, pyruvate oxidase, alcohol oxidase, bilirubin oxidase, uricase and the like. In many preferred embodiments in which the analyte of interest is glucose, the enzyme component of the redox reagent system is a glucose oxidase such as glucose oxidase or glucose dehydrogenase.

The second component of the redox reagent system is a mediator component consisting of one or more mediator formulations. A variety of different mediator agents are known in the art, examples of which are ferricyanide, phenazine ethosulfate, phenazine methosulfate, palenediamine, 1-methoxy-phenazine methosulfate, 2,6-dimethyl- 1,4-benzoquinone, 2,5-dichloro-1,4-benzoquinone, ferrocene derivatives, osmium bipyridyl complexes, ruthenium complexes, and the like. In certain embodiments where the analyte of interest is glucose and glucose oxidase or glucose dehydrogenase is an enzyme component, a particularly useful mediator is ferricyanide. Other reagents that may be present in the reaction zone include buffers such as citraconate, citrate, phosphate, "Good" buffers, and the like.

Redox reagent systems are generally present in anhydrous form. The amount of the various components can vary, and the amount of the enzyme component is typically about 0.1 to 10% by weight.

Way

According to the present invention, a method of using the electrochemical test strip to measure the concentration of an analyte in a physiological sample or a panel of an analyte is provided. Various different analytes can be detected using the test strips, in which case the most representative analytes include glucose, cholesterol, lactate, alcohols and the like. In many preferred embodiments, the present methods are used to determine glucose concentration in physiological samples. In principle, the method can be used to determine the concentration of analytes in a variety of different physiological samples (e.g., urine, tears, saliva, etc.), which in particular are of the blood or blood fraction, more particularly whole blood. It is suitable for measuring concentration.

In carrying out the method, the first step is to introduce an amount of physiological sample into one or more reaction zones of the electrochemical test strip described above. In general, the reaction zone nearest the edge of the strip is used first. In FIG. 5, the contact 28 of the coelectrochemical test strip 26 is shown to be inserted into the electrosensitizer 27, with the physiological sample applied to the final reaction zone 30 of the strip 26. .

The amount of physiological sample (eg blood) introduced into each reaction zone of the test strip can vary, but is generally about 0.1-10 μl, typically about 0.3-1.6 μl. The sample may be introduced into the reaction zone using any convenient protocol that is convenient for use, such as, for example, scanning the sample into the reaction zone, applying a gauze wick to the reaction zone, and the like.

In certain embodiments, the strip can be used for continuous detection analysis of the analyte. In these aspects, the sample is introduced into each reaction zone at different time points.

After the sample is applied to the reaction zone, electrochemical measurements are performed using a reference electrode and a working electrode. The electrochemical measurements performed may vary depending on the nature of the assay, and on the apparatus in which the electrochemical test strip is used, for example whether the assay is a calorimetry, amperometric or potentiometric measurement. In general, an electrochemical measurement is one in which a sample is introduced into a reaction region, and then, typically, a charge (electrometric measurement), a current (current measurement) or a potential (potentiometric measurement) is measured over a predetermined period. Methods of performing such electrochemical measurements are further described in US Pat. Nos. 4,224,125, 4,545,382, 5,266,179, WO97 / 18465, and WO99 / 49307, which are incorporated herein by reference. If two or more reaction zones are used at the same time, means are provided for electrically insulating each of the separate cells.

After detection of the electrochemical signal generated in the reaction zone as described above, the amount of analyte present in the sample introduced into the reaction zone is determined by correlating the amount of the analyte in the sample with the electrochemical signal. In making this derivation, the measured electrochemical signal is usually compared with a signal calculated from a series of control or standard values previously obtained and determined from this comparison. In many embodiments, the electrochemical signal measurement step and the analyte concentration derivation step as described above are designed to work with the test strip to obtain a value of the analyte concentration in the sample applied to the test strip. Performed automatically by the equipment. A representative reading device, ie meter, for automatically performing these steps is a device in which the user only needs to read the final analyte concentration result from the device after applying the sample to the reaction zone, see here. It is further described in co-pending US patent application Ser. No. 09 / 333,793, filed June 15, 1999, which is incorporated by reference.

If the reaction zones of the strip are used for different analyses, each reaction zone used can be separated from the rest of the strip before and after the use of the next reaction zone, if necessary, for example, to be cut out from the strip. Can be. This process is illustrated in FIGS. 3 and 4, in which a cut 25 is applied to the strip to remove each reaction zone used prior to use of the next reaction zone.

Systems and kits

Also provided by the present invention are systems and kits for use in carrying out the method. The systems and kits of the invention comprise at least the electrochemical test strips of the invention as described above. The system and kit may further comprise means for obtaining a physiological sample. For example, if the physiological sample is blood, the system and kit may further comprise means for obtaining a blood sample, for example a lance for poke a finger, a lance actuation means, and the like. In addition, the system and kit may comprise a control solution, eg, a glucose control solution containing a standardized concentration of glucose. In certain embodiments, the systems and kits also include metering equipment as described above for detecting electrochemical signals using electrodes after applying the sample and correlating the detected signal with the amount of analyte in the sample. Finally, the system and kit may include instructions for using the reagent test strips in determining the concentration of the analyte in the physiological sample. Such instructions may be present in one or more packaging, insert labels, containers contained in a kit, and the like.

The present invention provides a significant improvement in that each test strip can be used several times. As such, the present invention provides more efficient use of the strip and cost savings in terms of reducing waste. The present invention also provides an opportunity to test a panel of analytes using a single strip. As such, the present invention has made a significant contribution to the art.

All publications and patents cited in the present specification are herein incorporated by reference, as each individual publication and patent is specifically and individually indicated to be incorporated by reference. Since the citation of any publication was published before the filing date, it should not be construed as an admission that the present invention is not entitled to antedate such publication on the ground that it is a conventional invention.

Although the foregoing invention has been described in some detail by way of illustration and illustration for clarity of understanding, those skilled in the art will recognize that specific changes and modifications can be made to the invention without departing from the scope and spirit of the appended claims. You can easily see from.

The present invention provides a significant improvement in that each test strip can be used several times. As such, the present invention provides more efficient use of the strip and cost savings in terms of reducing waste. The present invention also provides an opportunity to test a panel of analytes using a single strip.

Claims (10)

(a) a plurality of reaction zones defined by opposing working and reference electrodes separated by a spacer layer; And (b) an electrochemical test strip comprising a reagent composition present in each of said reaction zones. The electrochemical test strip of claim 1, wherein the test strip comprises 2 to 25 reaction zones. The electrochemical test strip of claim 1, wherein each reaction zone contains the same reagent composition. The electrochemical test strip of claim 1, wherein the two or more reaction zones contain different reagent compositions. 5. The electrochemical test strip of claim 1, wherein each reaction zone has its own fluid entry path to provide for the interaction of fluid between the reaction zone and the external environment of the test strip. 6. . 6. The method of claim 1, wherein two or more reaction zones are combined to form a single entry passage to provide for the interaction of fluid between the reaction zone and the external environment of the test strip. An electrochemical test strip with fluid entry passages. The electrochemical test strip as claimed in claim 1, wherein the strip is present in the meter. (a) applying a physiological sample to an electrochemical test strip comprising opposing working electrodes separated by a spacer layer and a plurality of reaction zones defined by reference electrodes and reagent compositions present in each of the reaction zones; (b) detecting the electrical signal in the reaction zone using a metal electrode; And (c) correlating the detected electrical signal with the amount of analyte in the sample. (a) an electrochemical test strip comprising opposing working electrodes separated by a spacer layer and a plurality of reaction zones defined by reference electrodes and reagent compositions present in each of the reaction zones; And A kit for use in determining the concentration of an analyte in a physiological sample, comprising (b) (i) a means for obtaining a physiological sample and (ii) at least one of the analyte standards. (a) an electrochemical test strip comprising opposing working electrodes separated by a spacer layer and a plurality of reaction zones defined by reference electrodes and reagent compositions present in each of the reaction zones; And (b) a system for use in measuring the concentration of an analyte in a physiological sample, including a meter.