Patent Publication Number: US-8529741-B2

Title: System and methods for determining an analyte concentration incorporating a hematocrit correction

Description:
This application is a continuation of U.S. application No. 11/845,860, filed Aug. 28, 2007 now abandoned, claims to the benefit of U.S. Provisional Application No. 60/842,032, filed Sep. 5, 2006, all of which are incorporated herein by reference. 
    
    
     DESCRIPTION OF THE INVENTION 
     1. Field of the Invention 
     The present invention relates to the field of diagnostic testing systems for measuring the concentration of an analyte in a blood sample and, more particularly, to methods for measuring an analyte concentration that incorporates a hematocrit correction. 
     2. Background of the Invention 
     The present disclosure relates to a biosensor system for measuring an analyte in a bodily fluid, such as blood, wherein the system comprises a unique process and system for correcting inaccuracies in sample concentration measurements. For example, the present disclosure provides methods of correcting analyte concentration measurements of bodily fluids. 
     Electrochemical sensors have long been used to detect and/or measure the presence of substances in a fluid sample. In the most basic sense, electrochemical sensors comprise a reagent mixture containing at least an electron transfer agent (also referred to as an “electron mediator”) and an analyte specific bio-catalytic protein (e.g. a particular enzyme), and one or more electrodes. Such sensors rely on electron transfer between the electron mediator and the electrode surfaces and function by measuring electrochemical redox reactions. When used in an electrochemical biosensor system or device, the electron transfer reactions are transformed into an electrical signal that correlates to the concentration of the analyte being measured in the fluid sample. 
     The use of such electrochemical sensors to detect analytes in bodily fluids, such as blood or blood derived products, tears, urine, and saliva, has become important, and in some cases, vital to maintain the health of certain individuals. In the health care field, people such as diabetics, for example, have a need to monitor a particular constituent within their bodily fluids. A number of systems are available that allow people to test a body fluid, such as, blood, urine, or saliva, to conveniently monitor the level of a particular fluid constituent, such as, for example, cholesterol, proteins, and glucose. Patients suffering from diabetes, a disorder of the pancreas where insufficient insulin production prevents the proper digestion and utilization of sugar, have a need to carefully monitor their blood glucose levels on a daily basis. Routine testing and controlling blood glucose for people with diabetes can reduce their risk of serious damage to the eyes, nerves, and kidneys. 
     A number of systems permit people to conveniently monitor their blood glucose levels, and such systems typically include a test strip where the user applies a blood sample and a meter that “reads” the test strip to determine the glucose level in the blood sample. An exemplary electrochemical biosensor is described in U.S. Pat. No. 6,743,635 (&#39;635 patent) which is incorporated by reference herein in its entirety. The &#39;635 patent describes an electrochemical biosensor used to measure glucose level in a blood sample. The electrochemical biosensor system is comprised of a test strip and a meter. The test strip includes a sample chamber, a working electrode, a counter electrode, and fill-detect electrodes. A reagent layer is disposed in the sample chamber. The reagent layer contains an enzyme specific for glucose, such as, glucose oxidase, and a mediator, such as, potassium ferricyanide or ruthenium hexaamine. When a user applies a blood sample to the sample chamber on the test strip, the reagents react with the glucose in the blood sample and the meter applies a voltage to the electrodes to cause redox reactions. The meter measures the resulting current that flows between the working and counter electrodes and calculates the glucose level based on the current measurements. 
     Biosensors configured to measure a blood constituent may be affected by the presence of certain blood components that may undesirably affect the measurement and lead to inaccuracies in the detected signal. This inaccuracy may result in an inaccurate glucose reading, leaving the patient unaware of a potentially dangerous blood sugar level, for example. As one example, the particular blood hematocrit level (i.e., the percentage of the amount of blood that is occupied by red blood cells) can erroneously affect a resulting analyte concentration measurement. 
     Variations in a volume of red blood cells within blood can cause variations in glucose readings measured with disposable electrochemical test strips. Typically, a negative bias (i.e., lower calculated analyte concentration) is observed at high hematocrits, while a positive bias (i.e., higher calculated analyte concentration) is observed at low hematocrits. At high hematocrits, for example, the red blood cells may impede the reaction of enzymes and electrochemical mediators, reduce the rate of chemistry dissolution since there less plasma volume to solvate the chemical reactants, and slow diffusion of the mediator. These factors can result in a lower than expected glucose reading as less current is produced during the electrochemical process. Conversely, at low hematocrits, less red blood cells may affect the electrochemical reaction than expected, and a higher measured current can result. In addition, the blood sample resistance is also hematocrit dependent, which can affect voltage and/or current measurements. 
     Several strategies have been used to reduce or avoid hematocrit based variations on blood glucose readings as described in U.S. patent application Ser. No. 11/401,458 which is incorporated by reference herein in its entirety. For example, test strips have been designed to incorporate meshes to remove red blood cells from the samples, or have included various compounds or formulations designed to increase the viscosity of red blood cell and attenuate the effect of low hematocrit on concentration determinations. Further, biosensors have been configured to measure hematocrit by measuring optical variations after irradiating the blood sample with light, or measuring hematocrit based on a function of sample chamber fill time. These methods have the disadvantages of increasing the cost and complexity of test strips and may undesirably increase the time required to determine an accurate glucose measurement. 
     In addition, alternating current (AC) impedance methods have also been developed to measure electrochemical signals at frequencies independent of a hematocrit effect. Such methods suffer from the increased cost and complexity of advanced meters required for signal filtering and analysis. 
     An additional prior hematocrit correction scheme is described in U.S. Pat. No. 6,475,372. In that method, a two potential pulse sequence is employed to estimate an initial glucose concentration and determine a multiplicative hematocrit correction factor. A hematocrit correction factor is a particular numerical value or equation that is used to correct an initial concentration measurement, and may include determining the product of the initial measurement and the determined hematocrit correction factor. Data processing using this technique, however, is complicated because both a hematocrit correction factor and an estimated glucose concentration must be determined to establish the corrected glucose value. In addition, the time duration of the first step greatly increases the overall test time of the biosensor, which is undesirable from the user&#39;s perspective. 
     Accordingly, novel systems and methods for providing corrected analyte concentration measurements are desired that overcome the drawbacks of current biosensors and improve upon existing electrochemical biosensor technologies so that measurements are more accurate. 
     SUMMARY OF THE INVENTION 
     One embodiment is directed to a biosensor having a base layer including a first capillary disposed on the base layer configured to electrochemically determine a concentration of a first analyte in a blood sample, and wherein the first capillary includes a first set of at least one electrode. The biosensor also includes a second capillary disposed on the base layer configured to determine a value correlating to the hematocrit level of the blood sample, and wherein the second capillary includes a second set of at least one electrode. 
     Another embodiment of the invention is directed to a method for manufacturing a biosensor comprising at least partially forming a plurality of electrodes on a generally planar base layer. The method also includes forming a first capillary disposed on the base layer, and wherein the first capillary includes a first set of at least one electrode selected from the plurality of at least partially formed electrodes. Further, the method includes forming a second capillary on the base layer, and wherein the second capillary includes a second set of at least one electrode selected from the plurality of at least partially formed electrodes. 
     Another embodiment of the invention is directed to a reel for manufacturing biosensors comprising a generally planar base layer including a plurality of at least partially formed electrodes. Additionally, the reel includes a first capillary disposed on the base layer, and wherein the first capillary includes a first set of at least one electrode selected from the plurality of at least partially formed electrodes. The reel also includes forming a second capillary on the base layer, and wherein the second capillary includes a second set of at least one electrode selected from the plurality of at least partially formed electrodes. 
     Another embodiment of the invention is directed to a method of manufacturing a plurality of test strips for a biosensor comprising forming a reel containing a base layer. Moreover, the method includes forming a plurality of electrodes on the base layer, and partially forming a test strip, wherein the test strip includes a first capillary on the base layer including at least one of the plurality of electrodes and the test strip further includes a second capillary on the base layer including at least one of the plurality of electrodes. 
     Another embodiment of the invention is directed to a test card for quality control analysis of biosensors comprising a base layer, wherein the base layer includes a plurality of electrodes. Further, a plurality of partially formed test strips, wherein each test strip includes a first capillary on the base layer including at least one of the plurality of electrodes and each test strip further includes a second capillary on the base layer including at least one of the plurality of electrodes. 
     Additional objects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objects and advantages of the invention will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. 
     It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention, as claimed. 
     The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate several embodiments of the invention and together with the description, serve to explain the principles of the invention. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
       The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate several embodiments of the invention and together with the description, serve to explain the principles of the invention. 
         FIG. 1A  illustrates test media that can be produced using the methods of the present disclosure. 
         FIG. 1B  illustrates a test meter that can be used with test media produced according to the methods of the present disclosure. 
         FIG. 1C  illustrates a test meter that can be used with test media produced according to the methods of the present disclosure. 
         FIG. 2A  is a top plan view of a test strip according to an exemplary embodiment of the invention. 
         FIG. 2B  is a cross-sectional view of the test strip of  FIG. 2A , taken along line  2 B- 2 B. 
         FIG. 3A  shows a configuration of sample chambers on test strip  10  according to the methods of the present disclosure. 
         FIG. 3B  shows a configuration of sample chambers on test strip  10  according to the methods of the present disclosure. 
         FIG. 3C  shows a configuration of sample chambers on test strip  10  according to the methods of the present disclosure. 
         FIG. 4A  is a top view of a reel according to an exemplary disclosed embodiment of the invention. 
         FIG. 4B  is an enlarged tip view of a feature set on the reel of  FIG. 4A . 
         FIG. 5  is a top view of a test card according to a further illustrative embodiment of the invention. 
         FIG. 6  is a diagram of the manufacturing process before production testing according to a further illustrative embodiment of the invention. 
     
    
    
     DESCRIPTION OF THE EMBODIMENTS 
     Reference will now be made in detail to the exemplary embodiments of the invention, examples of which are illustrated in the accompanying drawings. Wherever possible, the same reference numbers will be used throughout the drawings to refer to the same or like parts. 
     In accordance with an exemplary embodiment, a biosensor manufacturing method is described. Many industries have a commercial need to monitor the concentration of particular constituents in a fluid. The oil refining industry, wineries, and the dairy industry are examples of industries where fluid testing is routine. In the health care field, people such as diabetics, for example, need to monitor various constituents within their bodily fluids using biosensors. A number of systems are available that allow people to test a body fluid (e.g., blood, urine, or saliva), to conveniently monitor the level of a particular fluid constituent, such as, for example, cholesterol, proteins or glucose. 
     For purposes of this disclosure, “distal” refers to the portion of a test strip further from the fluid source (i.e., closer to the meter) during normal use, and “proximal” refers to the portion closer to the fluid source (e.g., a finger tip with a drop of blood for a glucose test strip) during normal use. The test strip may include a plurality of sample chambers for receiving a user&#39;s fluid sample, such as, for example, a blood sample. The sample chambers and test strip of the present specification can be formed using materials and methods described in commonly owned U.S. Pat. No. 6,743,635, which is hereby incorporated by reference in its entirety. Accordingly, a sample chamber may include a first opening in the proximal end of the test strip and a second opening for venting the sample chamber. Each sample chamber may be dimensioned so as to be able to draw the blood sample in through the first opening and to hold the blood sample in the sample chamber by capillary action. The test strip can include a tapered section that is narrowest at the proximal end, or can include other indicia in order to make it easier for the user to locate the first opening and apply the blood sample. 
     A first set of electrodes, such as a working electrode and a counter (or in an exemplary embodiment, proximal) electrode, can be disposed in a first sample chamber optionally along with one or more fill-detect electrodes. A reagent layer is disposed in the first sample chamber and preferably contacts at least the working electrode. The reagent layer may include an enzyme, such as glucose oxidase or glucose dehydrogenase, and a mediator, such as potassium ferricyanide or ruthenium hexamine. The first sample chamber may be configured to permit determination of one or more analytes in a blood sample, such as, for example, glucose. A second set of electrodes may be disposed in a second sample chamber, such as, for example, a proximal electrode and a distal electrode. The electrodes may be spaced at a predetermined distance such that hematocrit may be determined by measurement of electrical impedance between the two electrodes in the second sample chamber. 
     The test strip has, near its distal end, a plurality of electrical contacts that are electrically connected to the electrodes via conductive traces. In addition, the test strip may also include a second plurality of electrical strip contacts near the distal end of the strip. The second plurality of electrical contacts can be arranged such that they provide, when the strip is inserted into the meter, a distinctly discernable lot code readable by the meter. In some embodiments, the electrical contacts may be at least partially covered with an at least partially conductive material to improve the wear properties of the electrical contacts. 
     An individual test strip may also include an embedded code relating to data associated with a lot of test strips, or data particular to that individual strip. The embedded information presents data readable by the meter signaling the meter&#39;s microprocessor to access and utilize a specific set of stored calibration parameters particular to test strips from a manufacturing lot to which the individual strip belongs, or to an individual test strip. The system may also include a check strip that the user may insert into the meter to check that the instrument is electrically calibrated and functioning properly. The readable code can be read as a signal to access data, such as calibration coefficients, from an on-board memory unit in the meter. 
     In order to save power, the meter may be battery powered and may stay in a low-power sleep mode when not in use. When the test strip is inserted into the meter, one or more electrical contacts on the test strip form electrical connections with one or more corresponding electrical contacts in the meter. The second plurality of electrical contacts may bridge a pair of electrical contacts in the meter, causing a current to flow through a portion of the second plurality of electrical contacts. The current flow through the second plurality of electrical contacts causes the meter to wake up and enter an active mode. The meter also reads the code information provided by the second plurality and can then identify, for example, the particular test to be performed or a confirmation of proper operating status. Calibration data pertaining to the strip lot, for either the analyte test or the hematocrit test, discussed below, can also be encoded or otherwise represented. In addition, based on the particular code information, the meter can also identify the inserted strip as either a test strip or a check strip. If the meter detects a check strip, it performs a check strip sequence. If the meter detects a test strip, it performs a test strip sequence. In the test strip sequence, the meter validates the working electrode, counter electrode, and, if included, the fill-detect electrodes, by confirming that there are no low-impedance paths between any of these electrodes. If the electrodes are valid, the meter indicates to the user that a sample may be applied to the test strip. The meter then applies a drop-detect voltage between any two suitable electrodes and detects a fluid sample, such as, a blood sample, by detecting a current flow between the working and proximal electrodes (i.e., a current flow through the blood sample as it bridges the working and proximal electrodes). To detect that an adequate sample is present in the sample chamber and that the blood sample has traversed the reagent layer and mixed with the chemical constituents in the reagent layer, the meter may apply a fill-detect voltage to the one or more fill-detect electrodes and measure any resulting current flow. If a resulting electrical property reaches a sufficient level within a predetermined period of time, the meter indicates to the user that adequate sample is present and has mixed with the reagent layer. 
     The meter can be programmed to wait for a predetermined period of time after initially detecting the blood sample to allow the blood sample to react with the reagent layer. Alternatively, the meter may be configured to immediately begin taking readings in sequence. During an exemplary fluid measurement sequence using amperometry, the meter applies an assay voltage between the working and proximal electrodes and takes one or more measurements of the resulting current flowing between the working and counter electrodes. The assay voltage is near the redox potential of the chemistry in the reagent layer, and the resulting current is related to the concentration of the particular constituent measured, such as, for example, the glucose level in a blood sample. Voltammetry and coulometry approaches, as known in the art, could also be employed. 
     In one example, the reagent layer may react with glucose in the blood sample in order to determine the particular glucose concentration. In one example, glucose oxidase or glucose dehydrogenase is used in the reagent layer. During a sample test, the glucose oxidase initiates a reaction that oxidizes the glucose to gluconic acid and reduces a mediator such as ferricyanide or ruthenium hexamine. When an appropriate voltage is applied to a working electrode relative to a counter electrode, the ferrocyanide is oxidized to ferricyanide, thereby generating a current that is related to the glucose concentration in the blood sample. 
     The test strip may also include a second sample chamber configured to permit determination of hematocrit. The meter can determine hematocrit by measuring the impedance of the blood sample in the second sample chamber by applying an appropriate voltage and/or current and reading suitable measurements to calculate an impedance value. The calculated impedance value correlates with hematocrit, which can vary and can affect glucose determination. 
     The meter can calculate the glucose level based on the measured current from the first sample chamber and, optionally, enhance that calculation based on the impedance value determined using the second sample chamber. This data along with other calibration data contained within the test strip may permit the meter to determine a glucose level and display the calculated glucose level to the user. 
     Electrodes positioned within the sample chamber may include a working electrode, a counter electrode, a fill-detect electrode, a proximal electrode, and a distal electrode. A reagent layer can be disposed in the first sample chamber and may cover at least a portion of the working electrode, which can also be disposed at least partially in the sample chamber. The reagent layer can include, for example, an enzyme, such as glucose oxidase or glucose dehydrogenase, and a mediator, such as potassium ferricyanide or ruthenium hexamine, to facilitate the detection of glucose in blood. It is contemplated that other reagents and/or other mediators can be used to facilitate detection of glucose and other constituents in blood and other body fluids. The reagent layer can also include other components, such as buffering materials (e.g., potassium phosphate), polymeric binders (e.g., hydroxypropyl-methyl-cellulose, sodium alginate, microcrystalline cellulose, polyethylene oxide, hydroxyethylcellulose, and/or polyvinyl alcohol), and surfactants (e.g., Triton X-100 or Surfynol 485). 
     As mentioned previously, biosensors may inaccurately measure a particular constituent level in blood due to unwanted effects of certain blood components on the method of measurement. For example, the hematocrit level (i.e., the percentage of blood occupied by red blood cells) in blood can erroneously affect a resulting analyte concentration measurement. Thus, it may be desirable to apply chemical additives and/or signal processing techniques as previously described, to reduce the sensitivity of the blood sample to hematocrit. Further, it may be desirable to separately measure hematocrit of a blood sample such that any analyte measurement can be adjusted to correct for hematocrit variations. In accordance with an exemplary embodiment of the present invention, a blood sample may be divided into at least two different regions on a biosensor and tested separately. For example, a blood sample may be diverted into a first sample chamber to undergo an electrochemical test, as described above, to determine, for example, the concentration of glucose within the sample. The blood sample may also be diverted into a second sample chamber to undergo a separate test, as discussed in detail below, to determine the hematocrit level of the blood sample. It is also contemplated that a third sample chamber may be used to perform another determination, such as, for example, a determination of blood sample temperature, a concentration of a second analyte, a second measurement of the first analyte concentration, and/or an on board control to perform a calibration step. In some embodiments, the second sample chamber may be configured to perform one or more determinations as described for the third sample chamber, as outlined in detail below. 
     In some embodiments, first and second sample chambers can be dimensioned and configured to draw a blood sample into the sample chambers via capillary action. Each sample chamber may also include one or more electrodes positioned within the sample chambers and configured to contact the blood sample. The first sample chamber may include reagents and electrodes configured to determine a blood glucose concentration. Hematocrit may be measured using the second sample chamber. For example, the second chamber may include a set of electrodes spaced apart at a predetermined distance, and hematocrit may be determined by measuring an impedance of the blood sample between the electrodes. The distance between the electrodes in the second sample chamber can be optimized for measuring hematocrit while the electrodes of the first sample chamber may be configured for glucose determination. 
     Hematocrit may be determined using any methods known in the art. For example, hematocrit may use electrical, optical, chemical, or any other suitable method. Optical methods may include reflective or transmission techniques. Electrical methods may include amperometric, voltametric, or coulometric. In some embodiments, hematocrit may be determined using an AC excitation, wherein an impedance measurement may be obtained using digital signal processing, analog processing, or a similar suitable technique. 
     To determine impedance, an AC signal can be applied across a set of electrodes in the second sample chamber. Impedance may include real or complex values, wherein effective, reactive, capacitive and/or resistive parameters may be associated with hematocrit. As explained by the Coulter principle, blood hematocrit can be derived from an impedance measurement obtained by applying an AC signal to the blood sample. More specifically, impedance Z R  can be measured from the blood sample by dividing the phasor voltage V, applied across the electrodes and dividing this value by the phasor current I r  passing through the electrodes and the blood sample. Thus, the impedance of the blood sample is: 
     
       
         
           
             
               Z 
               R 
             
             = 
             
               
                 V 
                 r 
               
               
                 I 
                 r 
               
             
           
         
       
     
     Following impedance measurement, hematocrit can be determined by applying the measured impedance value or multiple values at several different frequencies of excitation to an equation, an algorithm, a look-up chart, or any other suitable method. For example, an algorithm may correlate a glucose level with an electrical measurement value up to a threshold value, and above that threshold, a correction value correlated with hematocrit may be applied to any glucose determination. Once the value correlated to the hematocrit level within the blood sample is determined, the value may be used to modify the calculated glucose concentration such that an enhanced or corrected value of the concentration of glucose of the blood sample can be determined. Determining a glucose measurement and/or a hematocrit value may also require incorporating of one or more correction values, such as, for example, for variations in a temperature of a blood sample. 
     In accordance with another exemplary embodiment of the present invention, a test strip may further comprise a third sample chamber configured to permit determination of a third parameter associated with a blood sample. The third parameter to be measured may be selected from a group consisting of a temperature, a concentration of a second analyte, and an on-board control, as described in detail below. 
     In some embodiments, one or more sample chambers may be configured to receive a control solution. The control solution may be used to periodically test one or more functions of a meter. For example, a control solution may include a solution of known electrical properties and an electrical measurement of the solution may be performed by the meter. When the meter detects the use of a control solution, it can provide an operational check of both sample chambers functionality to verify the systems measurement integrity. The meter read-out may then be compared to the known glucose value of the solution to confirm that the meter is functioning to an appropriate accuracy. Any measurement of a control solution may be performed using one or more electrodes of the second sample chamber. In addition, data associated with a measurement of a control solution may be processed, stored and/or displayed using a meter differently to any data associated with a glucose measurement. Such different treatment of data associated with the control solution may permit a meter, or user, to distinguish a glucose measurement, or may permit exclusion of any control measurements when conducting any statistical analysis of glucose measurements. 
     The present disclosure provides a method for producing a diagnostic test strip  10 , as shown in  FIG. 1A . Test strip  10  of the present disclosure may be used with a suitable test meter  400 ,  408 , as shown in  FIGS. 1B and 1C , to detect or measure the concentration of one or more analytes. As shown in  FIG. 1A , test strip  10  are planar and elongated in design. However, test strip  10  may be provided in any suitable form including, for example, ribbons, tubes, tabs, discs, or any other suitable form. Furthermore, test strip  10  can be configured for use with a variety of suitable testing modalities, including electrochemical tests, photochemical tests, electro-chemiluminescent tests, and/or any other suitable testing modality. 
     Test meter  400 ,  408  may be selected from a variety of suitable test meter types. For example, as shown in  FIG. 1B , test meter  400  includes a vial  402  configured to store one or more test strips  10 . The operative components of test meter  400  may be contained in a meter cap  404 . Meter cap  404  may contain electrical meter components, can be packaged with test meter  400 , and can be configured to close and/or seal vial  402 . Alternatively, a test meter  408  can include a monitor unit separated from storage vial, as shown in  FIG. 1C . Any suitable test meter may be selected to provide a diagnostic test using test strip  10  produced according to the disclosed methods. 
     Test Strip Configuration 
     With reference to the drawings,  FIGS. 2A and 2B  show a test strip  10 , in accordance with an exemplary embodiment of the present invention. Test strip  10  preferably takes the form of a generally flat strip that extends from a proximal end  12  to a distal end  14 . Preferably, test strip  10  is sized for easy handling. For example, test strip  10  can measure approximately 35 mm long (i.e., from proximal end  12  to distal end  14 ) and approximately 9 mm wide. The strip, however, can be any convenient length and width. For example, a meter with automated test strip handling may utilize a test strip smaller than 9 mm wide. Additionally, proximal end  12  can be narrower than distal end  14  in order to provide facile visual recognition of the distal end. Thus, test strip  10  can include a tapered section  16 , in which the full width of test strip  10  tapers down to proximal end  12 , making proximal end  12  narrower than distal end  14 . As described in more detail below, the user applies the blood sample to an opening in proximal end  12  of test strip  10 . Thus, providing tapered section  16  in test strip  10 , and making proximal end  12  narrower than distal end  14 , assists the user in locating the opening where the blood sample is to be applied. Further, other visual means, such as indicia, notches, contours or the like are possible. 
     As shown in  FIG. 2B , test strip  10  can have a generally layered construction. Working upwardly from the bottom layer, test strip  10  can include a base layer  18  extending along the entire length of test strip  10 . Base layer  18  can be formed from an electrically insulating material and have a thickness sufficient to provide structural support to test strip  10 . For example, base layer  18  can be formed from a polyester (e.g., PET), acrylic, and/or other plastic material and be about 0.35 mm thick. 
     According to the illustrative embodiment, a conductive layer  20  is disposed on base layer  18 . Conductive layer  20  includes a plurality of electrodes disposed on base layer  18  near proximal end  12 , a plurality of electrical contacts disposed on base layer  18  near distal end  14 , and a plurality of conductive regions electrically connecting the electrodes to the electrical contacts. In the illustrative embodiment depicted in  FIG. 2A , the plurality of electrodes includes a working electrode  22 , a proximal electrode (or counting electrode)  24 , a distal electrode (or fill detect-anode)  28 , and a fill-detect electrode (or fill-detect cathode)  30 . Defined between proximal electrode  24  and distal electrode  28  is an electrically isolated region  26 , wherein the distance between electrodes  24  and  28  may be about 1 mm. The electrical contacts can correspondingly include a working electrode contact  32 , a proximal electrode contact  34 , a distal electrode contact  36 , and a fill-detect electrode contact  38 . The conductive regions can include a working electrode conductive region  40 , electrically connecting working electrode  22  to working electrode contact  32 , a proximal electrode conductive region  42 , electrically connecting proximal electrode  24  to proximal electrode contact  36 , a distal electrode conductive region  44  electrically connecting distal electrode  28  to distal electrode contact  36 , and a fill-detect electrode conductive region  46  electrically connecting fill-detect electrode  30  to fill-detect contact  38 . Further, the illustrative embodiment is depicted with conductive layer  20  including an auto-on conductor  48  disposed on base layer  18  near distal end  14 . 
     In addition, the present disclosure provides test strips  10  that include electrical contacts that are resistant to scratching or abrasion. Such test strips  10  can include conductive electrical contacts formed of two or more layers of conductive and/or semi-conductive material. Referring to  FIG. 2B , A first lower conductive layer  20  can include a conductive metal, ink, or paste. A second upper layer (not illustrated) can include a conductive ink or paste. Further, in some embodiments, the upper layer can have a resistance to abrasion that is greater than the lower layer. In addition, the second upper layer may have a thickness such that, even when scratched or abraded, the entire thickness of the conductive layer will not be removed, and the electrical contact will continue to function properly. Thus, such test strips  10  can include electrical contacts having material properties and dimensions such that, even when scratched or abraded, test strips  10  will continue to function properly. Further information relating to electrical contacts that are resistant to scratching or abrasion are described in U.S. patent application Ser. No. 11/458,298 which is incorporated by reference herein in its entirety. 
     The next layer in the illustrative test strip  10  is a dielectric spacer layer  64  disposed on conductive layer  20 . Dielectric spacer layer  64  is composed of an electrically insulating material, such as polyester (e.g., PET), acrylic, and/or other plastic material. Dielectric spacer layer  64  can be about 0.100 mm thick and covers portions of working electrode  22 , proximal electrode  24 , distal electrode  28 , fill-detect electrode  30 , and conductive regions  40 - 46 , but in the illustrative embodiment does not cover electrical contacts  32 - 38  or auto-on conductor  48 . For example, dielectric spacer layer  64  can cover substantially all of conductive layer  20  thereon, from a line just proximal of contacts  32  and  34  all the way to proximal end  12 , except for a first sample chamber  52  and a second sample chamber  58  extending from proximal end  12 . In this way, first sample chamber  52  can define an exposed portion  54  of working electrode  22 , an exposed portion  56  of proximal electrode  24 , and an exposed portion  62  of fill-detect electrode  30 . Second sample chamber  58  can define an exposed portion  59  of proximal electrode  24  and an exposed portion  60  of distal electrode  28 . In some embodiments, first sample chamber  52  may be configured to detect an analyte concentration in a blood sample and second sample chamber  58  may be configured to determine a hematocrit of the blood sample. The shape of sample chambers  52  and  58  may be achieved prior to application on the base layer. Alternatively sample chambers  52  and  58  may be formed subsequently, which may allow for tighter tolerances to be achieved in the formation of the sample chambers  52  and  58 . 
     A cover  72 , having a proximal end  74  and a distal end  76 , can be attached to dielectric spacer layer  64  via an adhesive layer  78 . Cover  72  can be composed of an electrically insulating material, such as polyester, and can have a thickness of about 0.1 mm. Additionally, cover  72  can be transparent. 
     Adhesive layer  78  can include a polyacrylic or other adhesive and have a thickness of about 0.013 mm. Adhesive layer  78  can consist of sections disposed on spacer layer  64  on opposite sides of first sample chamber  52 . A break  84  in adhesive layer  78  extends from a distal end  70  of first sample chamber  52  to an opening  86 . Cover  72  can be disposed on adhesive layer  78  such that its proximal end  74  is aligned with proximal end  12  and its distal end  76  is aligned with opening  86 . In this way, cover  72  covers first sample chamber  52  and break  84 . It is also contemplated that cover  72  may similarly cover second sample chamber  58 . 
     Proximal end  74  of cover  72  can extend from distal end  70  beyond proximal end  12  to create an overhang, as shown in  FIG. 2B . The overhang may be formed by extending cover  72  beyond proximal end  12  and/or by removing at least part of base layer  18  or other appropriate material under cover  72  to create a notch or similar structure. This overhang/notch configuration can aid in forming a hanging reservoir for a blood sample, via surface tension, to aid in providing a sufficient sample into first sample chamber  52  and second sample chamber  58 . It is also contemplated that various materials, surface coatings (e.g., hydrophilic and/or hydrophobic), or other structure protrusions and/or indentations at proximal end  12  may be used to form a suitable blood sample reservoir. 
     First sample chamber  52  and second sample chamber  58  may be configured to receive separate portions of a blood sample applied to test strip  10 . A proximal end  68  of first sample chamber  52  may define a first opening in first sample chamber  52 , through which the blood sample is introduced into first sample chamber  52 . At distal end  70  of first sample chamber  52 , break  84  may define a second opening in first sample chamber  52 , for venting first sample chamber  52  as a fluid sample enters first sample chamber  52 . First sample chamber  52  may be dimensioned such that a blood sample applied to its proximal end  68  may be drawn into first sample chamber  52  by capillary action, with break  84  venting first sample chamber  52  through opening  86 , as the blood sample enters. Moreover, first sample chamber  52  can advantageously be dimensioned so that the blood sample that enters first sample chamber  52  by capillary action is about 1 micro-liter or less. For example, first sample chamber  52  can have a length (i.e., from proximal end  12  to distal end  70 ) of about 0.140 inches, a width of about 0.060 inches, and a height (which can be substantially defined by the thickness of dielectric spacer layer  64 ) of about 0.005 inches. Other dimensions could be used, however. 
     Proximal end  12  of second sample chamber  58  may define a first opening in second sample chamber  58 , through which the blood sample is introduced into second sample chamber  58 . Second sample chamber  58  may be dimensioned such that a blood sample applied to its proximal end may be drawn into second sample chamber  58  by capillary action. Additionally, second sample chamber  58  can advantageously be dimensioned so that the blood sample that enters second sample chamber  58  by capillary action is about 0.5 micro-liters or less. 
     In some embodiments, a secondary sample chamber may be configured for operation with a continuous glucose monitoring system (not shown). Such a system may include systems and/or devices configured to automatically monitor a patient&#39;s glucose level. Such systems may periodically sample body fluid containing cellular or biological matter that may affect a glucose determination. Such systems may also benefit by using a secondary sample chamber configured to determine hematocrit, or a similar measurement, using one of more of the methods described here. 
       FIGS. 3A ,  3 B and  3 C show various illustrative embodiments of test strip  10 . Specifically, first sample chamber  52  and second sample chamber  58  may be variously configured in test strip  10 . As shown in  FIG. 3A , first sample chamber  52  and second sample chamber  58  may be arranged in a bifurcated configuration on test strip  10 , wherein first sample chamber  52  may be fluidly connected to a blood reservoir  63  and second sample chamber  58  may be fluidly connected to blood reservoir  63 . Such a configuration may permit fluid to flow from blood reservoir  63  into first sample chamber  52  (via an inlet) and second sample chamber  58  (via a second inlet) at appropriate flow rates. It is contemplated that first sample chamber  52  and second sample chamber  58  may also share a common inlet. 
       FIG. 3B  shows test strip  10  according to another illustrative embodiment. Test strip  10  may include first sample chamber  52  fluidly connected to second sample chamber  58  such that a blood sample may flow from one sample chamber into another sample chamber. For example, test strip  10  may be configured such that a blood sample applied to proximal end  12  flows into an inlet of second sample chamber  58 . The sample may, through capillary action, flow through second sample chamber  58  to an outlet of second sample chamber  58 , which may be fluidly connected to an inlet of first sample chamber  52 . Again, through capillary action, the blood sample may flow through first sample chamber  52 . Such a configuration may permit a blood sample to flow into second sample chamber  58  wherein hematocrit may be determined, and flow into first sample chamber  52  wherein a glucose concentration may be determined. It is also contemplated that sample chambers  52 ,  58  may be configured such that blood flows from the blood sample into first sample chamber  52 , and from first sample chamber  52  into second sample chamber  58 . 
       FIG. 3C  shows test strip  10  according to another illustrative embodiment, wherein test strip  10  may include a third sample chamber  61 . Third sample chamber  61  may be configured to permit determination of a third parameter associated with the blood sample, such as, for example, a blood sample temperature, a concentration of a second analyte within the blood sample, a second measurement associated with the first analyte, or any suitable measurement. Third sample chamber  61  may include one or more different or shared components associated with first sample chamber  52  and second sample chamber  58 , such as, for example, one or more electrodes, or reagent layers. 
     In some embodiments, third sample chamber  61  may be configured to provide an on-board control, wherein a function and/or calibration of test strip  10  may be conducted to at least partially confirm the accuracy of a measurement associated with test strip  10 . The third sample chamber  61  may be fluidly connected to first and/or second sample chambers  52 ,  58 . These and other configurations of multiple sample chambers within test strip  10  are contemplated within the scope of the present invention. 
     As shown in  FIG. 2B , a reagent layer  90  is disposed in first sample chamber  52 , wherein reagent layer  90  may include one or more chemical constituents to enable the level of glucose in the blood sample to be determined electrochemically. Thus, reagent layer  90  may include an enzyme specific for glucose and a mediator, as described above. In addition, reagent layer  90  may also include other components, buffering materials (e.g., potassium phosphate), polymeric binders (e.g., hydroxypropyl-methyl-cellulose, sodium alginate, microcrystalline cellulose, polyethylene oxide, hydroxyethylcellulose, and/or polyvinyl alcohol), and surfactants (e.g., Triton X-100 or Surfynol 485). 
     As depicted in  FIG. 2B , the arrangement of the various layers in illustrative test strip  10  can result in test strip  10  having different thicknesses in different sections. In particular, among the layers above base layer  18 , much of the thickness of test strip  10  can come from the thickness of spacer layer  64 . Thus, the edge of spacer layer  64  that is closest to distal end  14  can define a shoulder  92  in test strip  10 . Shoulder  92  can define a thin section  94  of test strip  10 , extending between shoulder  92  and distal end  14 , and a thick section  96 , extending between shoulder  92  and proximal end  12 . The elements of test strip  10  used to electrically connect it to the meter, namely, electrical contacts  32 - 38  and auto-on conductor  48 , can all be located in thin section  94 . Accordingly, the connector in the meter can be sized and configured to receive thin section  94  but not thick section  96 , as described in more detail below. This can beneficially cue the user to insert the correct end, i.e., distal end  14  in thin section  94 , and can prevent the user from inserting the wrong end, i.e., proximal end  12  in thick section  96 , into the meter. Although  FIGS. 2A and 2B  illustrate an illustrative embodiment of test strip  10 , other configurations, chemical compositions and electrode arrangements could be used. 
     As depicted in  FIG. 2A  fill-detect electrode  30  can function with working electrode  22  to perform a fill-detect feature, as previously described. Further, working electrode  22  may operate in conjunction with proximal electrode  24  to detection of a constituent of a sample in first sample chamber  52 , as described above. Other configurations of electrodes on test strip  10  are possible, such as, for example, multiple fill-detect electrodes and multiple working electrodes. 
     As depicted in the  FIG. 2B , fill-detect electrode  30  is advantageously located on the distal side of reagent layer  90 . In this arrangement, the sample introduced into first sample chamber  52  will have traversed reagent layer  90  before reaching fill-detect electrode  30 . This arrangement beneficially allows the fill-detect electrode  30  to indicate not only whether sufficient blood sample is present in first sample chamber  52 , but also when, concomitantly, the blood sample has sufficiently mixed with the chemical constituents of reagent layer  90 . 
     Test Strip Array Configuration 
     Test strips can be manufactured by forming a plurality of strips in an array along a reel or web of substrate material. The term “reel” or “web” as used herein applies to continuous webs of indeterminate length, or to sheets of determinate length. The individual strips, after being formed, can be separated during later stages of manufacturing. An illustrative embodiment of a batch process of this type is described below. First, an illustrative test strip array configuration is described. 
       FIG. 4A  shows a series of traces  80  formed in a substrate material coated with a conductive layer. Traces  80 , formed in the exemplary embodiment by laser ablation, partially form the conductive layers of two rows of ten test strips as shown. In the exemplary embodiment depicted, proximal ends  12  of the two rows of test strips are in juxtaposition in the center of a reel  100 . Distal ends  14  of test strips  10  are arranged at the periphery of reel  100 . It is also contemplated that the proximal ends  12  and distal ends  14  of test strips  10  can be arranged in the center of reel  100 . Alternatively, the two distal ends  14  of test strips  10  can be arranged in the center of reel  100 . The lateral spacing of test strips  10  may be designed to allow a single cut to separate two adjacent test strips. The separation of test strip  10  from reel  100  can electrically isolate one or more conductive components of the separated test strip  10 . 
     As depicted in  FIG. 4A , trace  80  for an individual test strip forms a plurality of conductive components; e.g., electrodes, conduction regions and electrode contacts. Trace  80  is comprised of individual cuts made by a laser following a specific trajectory, or vector. A vector can be linear or curvilinear, and define spaces between conductive components that are electrically isolating. Generally a vector is a continuous cut made by the laser beam. 
     The conductive components can be partially or entirely defined by ablated regions, or laser vectors, formed in the conductive layer. The vectors may only partially electrically isolate the conductive component, as the component can remain electrically connected to other components following laser ablation. The electrical isolation of the conductive components can be achieved following “singulation,” when individual test strips are separated from reel or web  100 . It is also contemplated that other conductive components may be electrically isolated during the laser ablation process. For example, fill detect electrodes may be isolated with the addition of one or more vectors. 
       FIG. 4A  also includes registration points  102  at the distal end  14  of each test strip on reel  100 . Registration points  102  assist the alignment of the layers during lamination, punching, etching, scoring, drilling, heating, compression, molding, printing, and/or other manufacturing processes. It is further contemplated that registration points  102  may be located at locations other than the distal end  14  of each test strip trace  80  on reel  100 . High quality manufacturing may require additional registration points  102  to ensure adequate alignment of laminate layers and/or other manufacturing processes, such as, for example, laser ablation of conductive components, reagent deposition, singulation, etc. It is contemplated that registration points  102  may be separated by less than 500 mm and may be less than 10 mm wide. 
       FIG. 5  shows a “test card”  104  separated from reel  100 . Test card  104  can contain a plurality of test strips  10  or traces  80 , and a plurality of conductive components. In the preferred embodiment test card  104  can contain between 6 and 12 test strips  10  or traces  80 . In other embodiments, test card  104  can contain a plurality of test strips  10  or traces  80 . In the illustrated embodiment, test card  104  can include a lateral array of test strips  10  or traces  80 . In other embodiments, test card  104  can include an array or arrays of test strips  10  or traces  80  in longitudinal and/or lateral configurations. It is further contemplated that test strips  10  or traces  80  may be in any arrangement on reel  100  suitable for manufacturing. 
     Test card  104  contains a plurality of conductive components. Some conductive components can be electrically isolated when test card  104  is removed from reel  100 . As shown in  FIG. 5 , working electrode  22  is electrically isolated. Other embodiments could include additional electrically isolated conductive components not shown in  FIG. 5 . It may be possible to analyze properties of the electrically isolated conductive components to assess the quality of the manufacturing process. The efficiency of the quality assessment process can be increased by testing at least one of the plurality of electrically isolated conductive components. 
     Batch Manufacturing of Test Strips 
     Test strip  10  may be manufactured using any suitable manufacturing methods. For example, one or more conductive components may be manufactured using laser ablation employing projected masks or raster scanning methods, screen printing, insert injection molding, and any other suitable techniques. One or more sample chambers, or capillaries, may be formed using a spacer, dielectric build-up, injection molded, laser ablation, or other suitable method. One illustrative embodiment for manufacturing test strip  10  will now be described in detail. 
       FIGS. 4A through 6  illustrate an exemplary method of manufacturing test strips. Although these figures shows steps for manufacturing test strip  10 , as shown in  FIGS. 4A through 6 , it is to be understood that similar steps can be used to manufacture test strips having other configurations. 
     With reference to  FIG. 5 , a plurality of test strips  10  can be produced by forming a structure  120  that includes a plurality of test strip traces  122  on reel  100 . Test strip traces  122  include a plurality of traces  80 , and can be arranged in an array that includes a plurality of rows. Each row  124  can include a plurality of test strip traces  122 . 
     The separation process can also be used to electrically isolate conductive components of test strip  10 . Laser ablation of the conductive layer may not electrically isolate certain conductive components. The non-isolated conductive components may be isolated by the separation process whereby test strips are separated from reel  100 . The separation process may sever the electrical connection, isolating the conductive component. Separating test strip  10  can electrically isolate the counting electrode  24 , fill detect-anode  28  and fill-detect cathode  30 . The separation process can complete the electrical isolation of conductive components by selectively separating conductive components. 
     Further, the separation process can provide some or all of the shape of the perimeter of test strips  10 . For example, the tapered shape of tapered sections  16  of test strips  10  can be formed during this punching process. Next, a slitting process can be used to separate test strip traces  122  in each row  124  into individual test strips  10 . The separation process may include stamping, slitting, scoring and breaking, or any suitable method to separate test strip  10  and/or card  104  from reel  100 . 
       FIGS. 4A and 4B  show only one test strip trace  122  (either partially or completely fabricated), in order to illustrate various steps in a preferred method for forming test strip traces  122 . In this exemplary approach, test strip traces  122  in integrated structure  120  are all formed on a sheet of material that serves as base layer  18  in the finished test strips  10 . The other components in the finished test strips  10  are then built up layer-by-layer on top of base layer  18  to form test strip traces  122 . In each of  FIGS. 4A and 4B , the outer shape of test strip  10  that would be formed in the overall manufacturing process is shown as a dotted line. 
     The exemplary manufacturing process employs base layer  18  covered by conductive layer  20 . Conductive layer  20  and base layer  18  can be in the form of a reel, ribbon, continuous web, sheet, or other similar structure. Conductive layer  20  can include any suitable conductive or semi-conductor material, such as palladium, gold, platinum, silver, iridium, carbon, indium tin oxide, indium zinc oxide, copper, aluminum, gallium, iron, mercury amalgams, tantalum, titanium, zirconium, nickel, osmium, rhenium, rhodium palladium, an organometallic, and/or other conductive or semi-conductor materials known in the art. Conductive layer  20  can be formed by sputtering, vapor deposition, screen printing or any suitable manufacturing method. For example, one or more electrodes may be at least partially formed by sputtering, evaporation, electroplating, ultrasonic spraying, pressure spraying, direct writing, shadow mask lithography, lift-off lithography, or laser ablation. Also, the conductive material can be any suitable thickness and can be bonded to base layer  18  by any suitable means. 
     As shown in  FIG. 2A , conductive layer  20  can include working electrode  22 , proximal electrode  24 , distal electrode  28 , and fill-detect cathode  30 . Trace  80  can be formed by laser ablation where laser ablation can include any device suitable for removal of the conductive layer in appropriate time and with appropriate precision and accuracy. Various types of lasers can be used for sensor fabrication, such as, for example, solid-state lasers (e.g. Nd:YAG and titanium sapphire), copper vapor lasers, diode lasers, carbon dioxide lasers and excimer lasers. Such lasers may be capable of generating a variety of wavelengths in the ultraviolet, visible and infrared regions. For example, excimer laser provides wavelength of 248 nm, a fundamental Nd:YAG laser gives 1064 nm, a frequency tripled Nd:YAG wavelength is at 355 nm and a Ti:sapphire laser is at approximately 800 nm. The power output of these lasers may vary and is usually in range 10-100 watts. 
     The laser ablation process can include a laser system. The laser system can include a laser source. The laser system can further include means to define trace  80 , such as, for example, a focused beam, projected mask or other suitable technique. The use of a focused laser beam can include a device capable of rapid and accurate controlled movement to move the focused laser beam relative to conductive layer  20 . The use of a mask can involve a laser beam passing through the mask to selectively ablate specific regions of conductive layer  20 . A single mask can define test strip trace  80 , or multiple masks may be required to form test strip trace  80 . To form trace  80 , the laser system can move relative to conductive layer  20 . Specifically, the laser system, conductive layer  20 , or both the laser system and conductive layer  20  may move to allow formation trace  80  by laser ablation. Exemplary devices available for such ablation techniques include Microline Laser system available from LPKF Laser Electronic GmbH (Garbsen, Germany) and laser micro machining systems from Exitech, Ltd (Oxford, United Kingdom). 
     In the next step, dielectric spacer layer  64  can be applied to conductive layer  20 , as illustrated in  FIG. 2B , Spacer layer  64  can be applied to conductive layer  20  in a number of different ways. In an exemplary approach, spacer layer  64  is provided as a sheet or web large enough and appropriately shaped to cover multiple test strip traces  80 . In this approach, the underside of spacer layer  64  can be coated with an adhesive to facilitate attachment to conductive layer  20 . Portions of the upper surface of spacer layer  64  can also be coated with an adhesive in order to provide adhesive layer  78  in each of test strips  10 . Various sample chambers can be cut, formed or punched out of spacer layer  64  to shape it before, during or after the application of spacer layer  64  to conductive layer  20 . In addition, spacer layer  64  can include adhesive sections and can include a break for each test strip trace. Spacer layer  64  is then positioned over conductive layer  20 , as shown in  FIG. 2B , and laminated to conductive layer  20 . When spacer layer  64  is appropriately positioned on conductive layer  20 , exposed electrode portions  54 - 62  are accessible through sample chambers  52  and  58 . Similarly, spacer layer  64  leaves contacts  32 - 38  and auto-on conductor  48  exposed after lamination. 
     Alternatively, spacer layer  64  could be applied in other ways. For example, spacer layer  64  can be injection molded onto base layer  18  and a substrate. Spacer layer  64  could also be built up on dielectric layer  50  by screen-printing successive layers of a dielectric material to an appropriate thickness, e.g., about 0.005 inches. A preferred dielectric material comprises a mixture of silicone and acrylic compounds, such as the “Membrane Switch Composition 5018” available from E.I. DuPont de Nemours &amp; Co., Wilmington, Del. Other materials could be used, however. 
     Additionally, sample chambers can be formed after application of spacer layer  64  on top of base layer  18  and conductive layer  20  via the aforementioned laser ablation process. This process allows for the removal of the conductive layer within sample chambers. 
     Reagent layer  90  can then be applied to each test strip structure. In an illustrative approach, reagent layer  90  is applied by dispensing a formulation onto exposed portion  54  of working electrode  22  and letting it dry to form reagent layer  90 . Alternatively, other methods, such as screen-printing, spray deposition, piezo and ink jet printing, can be used to apply the composition used to form reagent layer  90 . 
     An exemplary formulation contains 250 mM potassium phosphate at pH 6.75, 175-190 mM ruthenium hexamine, 5000 U/mL glucose dehydrogenase, 0.5-2.0% polyethylene oxide, 0.025-0.20% NATROSOL 250M (hydroxyethylcellulose), 0.675-2.5% Avicel (microcrystalline cellulose), 0.05-0.20% Triton-X surfactant and 2.5-5.0% trehalose. In some embodiments, various constituents may be added to reagent layer  90  to at least partially reduce a hematocrit bias of any measurement. For example, various polymers, molecules, and/or compounds may be added to reagent layer  90  to reduce cell migration and hence may increase the accuracy of a measurement based on an electrochemical reaction. Also, one or more conductive components may be coated with a surface layer (not shown) to at least partially restrict cell migration onto the one or more conductive components. These and other techniques known in the art may be used to reduce hematocrit bias from any measurement. 
     Cover  72  can then be attached to adhesive layer  78 . Cover  72  may be large enough to cover multiple test strip traces  122 . Attaching cover  72  can complete the formation of the plurality of test strip traces  122 . The plurality of test strip traces  122  can then be separated from each other to form a plurality of test strips  10 , as described above. 
     Quality Control Testing of Test Strips 
       FIG. 6  shows a further illustrative embodiment of a test strip manufacturing method. The manufacturing method utilizes a web  200  containing conductive layer  20  and base layer  18 . Conductive layer  20  and base layer  18  can be any suitable material. Web  200  can be any dimension suitable for production of test strips  10 . Web  200  is passed through any suitable device and ablated by process  300 . 
     Ablation  300  can include any suitable ablation process capable of forming conductive components in conductive layer  20 . In the illustrative embodiment, ablation  300  is achieved by laser ablation. The ablation process may not electrically isolate all conductive components. For example, counting electrode  24  may not be isolated by laser ablation but can be isolated by subsequent separation from web  200 . In the illustrative embodiment, working electrode  22  is electrically isolated during ablation process  300 . The proximal electrode  24 , distal electrode  28  and fill-detect cathode  30  may not be electrically isolated during ablation process  300 . Specifically, subsequent separation process can electrically isolate the proximal electrode  24 , distal electrode  28  and fill-detect cathode  30 . 
     Web  200  can be passed through any suitable ablation device at speeds sufficient to produce an appropriate rate of test strip production. The ablation process can be sufficiently rapid to allow the continuous movement of web  200  through the laser ablation device. Alternatively, web  200  can be passed through the ablation device in a non-continuous (i.e., start-and-stop) manner. 
     The properties of the conductive components formed by ablation process  300  can be analyzed during or following ablation process  300 . Analysis of ablation process  300  can include optical, chemical, electrical or any other suitable analysis means. The analysis can monitor the entire ablation process, or part of the ablation process. For example, the analysis can include monitoring vector formation to ensure the dimensions of the formed vector are within predetermined tolerance ranges. 
     Quality control analysis, which can be performed during or upon completion of the manufacturing process, can also include monitoring the effectiveness and/or efficiency of the vector formation process. In particular, the width of the resulting vectors can be monitored to ensure acceptable accuracy and precision of the cuts in conductive layer  20 . For example, the quality of the laser ablation process can be analyzed by monitoring the surface of conductive layer  20  and/or base layer  18  following ablation. Partial ablation of base layer  18  can indicate that the laser power is set too high or the beam is traveling too slowly. By contrast, a partially ablated conductive layer may indicate insufficient laser power or that the beam is traveling too quickly. Incomplete ablation of gaps may result in the formation of vectors that are not electrically isolating between conductive components. 
     In the illustrative embodiment, the dimensions of working electrode  22  can be analyzed to determine the quality of the manufacturing process. For example optical analysis (not shown) can monitor the width of working electrode  22  to ensure sufficient accuracy of ablation process  300 . Further, the alignment of working electrode  22  relative to registration points  102  can be monitored. Optical analysis can be performed by using VisionPro system from Cognex Vision Systems (Natick, Mass.). 
     As described above, the ablation process produces an array of test strips  202  on web  200 . Following formation of test strip array  202  and corresponding conductive components, dielectric spacer layer  64  is laminated to conductive layer  20 . A spacer lamination process  302  can include registration points  102  to correctly align spacer layer  64  with conductive layer  20 . Spacer layer  64  may contain registration points  102  corresponding to registration points  102  of test strip array  202 . Spacer lamination process may output a three layer laminate  204 . 
     A test card  206  may be separated from three layer laminate  204 . The separation may be achieved using punching, slitting, cutting, or any other appropriate process. Test card  206  can be analyzed by a test card analysis process  306  to test the quality of any previous manufacturing process. Test card analysis process  306  can include optical, electrical, chemical or any other suitable means for testing test card  206 . In an illustrative embodiment, the electrical properties of working electrode  22  can be tested. At least one of the plurality of working electrodes  22  of test card  206  can be analyzed for electrochemical and surface properties. For example, chronoamperometry can be used to test working electrode  22 . Chronoamperometry is an electrochemical technique that uses a voltage signal for excitation and measures current generated as a result of the excitation as a function of time. 
     Further, test card analysis process  306  may include measuring the width of space  26  between proximal electrode  24  and distal electrode  28  for accuracy. Additionally, a test card  104  may comprise test strips  10  in which sample chambers  52  and  58  have been formed, as discussed above. Under such circumstances test card analysis process  306  may include testing at least one of sample chambers  52  and  58  to determine if they have the dimensions that fit within predetermined tolerances, for example. 
     The results of test card analysis process  306  can be compared to previous manufacturing process. Alternatively, the results of test card analysis process  306  may be compared to modeled or simulated results using computational methods. The results can be used to ensure high-quality manufacturing processes. Deviation from acceptable or expected results may require altering upstream manufacturing processes, or altering downstream manufacturing processes to address the deviations. Following acceptance of the results of test card analysis process  306 , the quality of upstream manufacturing processes can be confirmed. 
     Following a satisfactory feedback  308  from test card analysis process  306 , the chemistry can be applied to three-layer laminate  204  by a chemistry application process  310 . A resulting laminate  208  can contain any appropriate reagent suitable for the specific test strip. A reagent application process  310  can include any appropriate process. In the preferred embodiment, quality control testing is not performed following reagent application process  310 . In other embodiments, quality control analysis can be conducted following reagent application process  310 . For example, quality control analysis can monitor the effectiveness of the chemistry application. Specifically, optical analysis may be required to determine the extent of reagent covering working electrode  22  and/or counter electrode  24 . Alternatively, any previous or upstream manufacturing process can be tested following formation of laminate  208 . 
     Following reagent application process  310 , cover  72  can be applied to laminate  208  using any appropriate cover application process  312 . Cover  72  may be centered on laminate  208 . The resulting laminate  210  can be tested to ensure the quality of the cover application process  312 . For example, optical means can be used to monitor the alignment of cover  72  to laminate  210 . Alternatively, laminate  210  can be tested to ensure the quality of any upstream manufacturing process as described previously. Following cover application process  312 , laminate  210  can be moved to a production testing  314 . 
     The manufacturing process can be halted at any stage based upon the results of the quality control testing during manufacturing or production. Alternatively, one or more manufacturing processes can be adjusted based on the results of the quality control analysis. Quality control tests can be conducted in real time, and/or may include analysis of test cards removed from the production line. If the quality control testing is performed on test cards taken out of the production line, any production of the same lot or batch can be intercepted in the manufacturing process downstream of the quality control testing. Test card  206  can contain addressable information, identifying where the test card was removed from the production line. Consequently any deviations from appropriate manufacturing quality can be isolated to specific regions of the production line. 
     Conclusion 
     In summary, determining hematocrit levels by measuring the impedance of a blood sample in a separate sample chamber has a number of advantages. It can be applied to many biosensors, not just oxidation-based glucose sensors. It has a high degree of accuracy and precision in regards to measuring and correcting for hematocrit since the presence of a separate chamber allows for optimizing the distance between the proximal and distal electrode specifically for measuring impedance of a blood sample. Further, since the impedance measurement can occur concurrently with the electrochemical measurement in another sample chamber, the amount of test time can be kept to a minimum. 
     While various test strip structures and manufacturing methods are described as possible candidates for use to measure HCT, they are not intended to be limiting of the claimed invention. Unless expressly noted, the particular test strip structures and manufacturing methods are listed merely as examples and are not intended to be limiting of the invention as claimed. Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims.