Biosensor with peroxidase enzyme

Abstract

An improved biosensor having at least a first working electrode and a first electrode material disposed on the first working electrode. The first electrode material is a mixture made by combining at least one enzyme where the at least one enzyme is a capable of reacting with the analyte to be measured, a redox mediator capable of reacting with a product of an enzymatic reaction or a series of enzymatic reactions involving the at least one enzyme, a peroxidase capable of catalyzing a reaction involving the redox mediator where the redox mediator is oxidized, a binder and a surfactant.

1. A disposable electrode strip for measuring an analyte in a fluid sample said strip comprising:
 a laminated strip having a first strip end, a second strip and and a vent opening spaced from said first strip and, said laminated strip comprising a base layer with at least two electrodes delineated thereon, a reagent holding layer carried on said base layer, said reagent holding layer having at least two cutouts, a channel forming layer carried on said reagent holding layer, and a cover;  
 an enclosed channel between said first strip end and said vent opening, said enclosed channel containing said at least two cutouts;  
 a first reagent disposed in a first cutout of said at least two cutouts forming a reference electrode, said first reagent comprising a reference electrode material selected from the group consisting of silver chloride when said reference electrode is silver and a mixture made by combining at least a redox mediator and at least one binder when said reference electrode is selected from the group consisting of gold, gold/tin oxide, palladium, platinum and carbon composition;  
 a second reagent disposed in a second cutout of said at least two cutouts forming a first working electrode, said second reagent comprising a redox mediator, at least one binder, at least one enzyme that is a substrate of said analyte and a peroxidase capable of catalyzing a reaction involving said redox mediator wherein said redox mediator is oxidized wherein said second reagent is water soluble; and  
 conductive contacts at said second strip end and insulated from said enclosed channel.  
 
     
     
       2. The electrode strip of  claim 1  further comprising a third cutout and a third reagent disposed in said third cutout forming a second working electrode wherein said third reagent comprises said redox mediator and said at least one binder wherein said third reagent is water soluble. 
     
     
       3. The electrode strip of  claim 2  wherein said third reagent further includes said at least one enzyme, a substrate of said at least one enzyme and a peroxidase. 
     
     
       4. The electrode strip of  claim 2  wherein said first reagent, said second reagent and said third reagent are made from a mixture having starting components comprising about 1 wt % to about 6.5 wt % of said redox mediator, about 1 wt % of binder, and about 0.02 wt % of said surfactant in water. 
     
     
       5. The electrode strip of  claim 4  wherein said first reagent, said second reagent and said third reagent further includes about 0.05 wt % to about 0.1 wt % of an antioxidant. 
     
     
       6. The electrode strip of  claim 4  wherein said second reagent is made from a mixture having starting components in water comprising about 2 wt % of potassium ferrocyanide, about 1 wt % of methyl cellulose, about 0.02 wt % of said t-octylphenoxypolyethoxyethanol, about 0.5 wt % of glucose oxidase, and about 0.5 wt % of soybean peroxidase. 
     
     
       7. The electrode strip of  claim 4  wherein said second reagent is made from a mixture having starting components in water comprising about 5 wt % of potassium ferrocyanide, about 1 wt % of methyl cellulose, about 0.02 wt % of t-octylphenoxypolyethoxyethanol, about 2 wt % of cholesterol oxidase, about 1 wt % of cholesterol esterase, and about 0.5 wt % of soybean peroxidase. 
     
     
       8. The electrode strip of  claim 4  wherein said second reagent is made from a mixture having starting components in water comprising about 2 wt % of potassium ferrocyanide, about 1 wt % of methyl cellulose, about 0.02 of t-octylphenoxypolyethoxyethanol, about 2 wt % of creatine amidinohydrolase, about 0.5 wt % of sarcosine oxidase, and about 0.5 of soybean peroxidase, and wherein said third reagent is made from a mixture having starting components in water comprising about 2 wt % of said potassium ferrocyanide, about 1 wt % of said methyl cellulose, about 0.02 wt % of said t-octylphenoxypolethoxyethanol, about 2 wt % of said creatine amidinohydrolase, about 0.4 wt % of creatinine amidohydrolase, about 0.5 wt % of said sarcosine oxidase, and about 0.5 wt % of said soybean peroxidase. 
     
     
       9. The electrode strip of  claim 8  wherein said first reagent and said second reagent further includes about 0.05 wt % of an antioxidant. 
     
     
       10. The electrode strip of  claim 2  wherein the surface area of said first working electrode is substantially the same size as the surface area of said second working electrode. 
     
     
       11. The electrode strip of  claim 1  wherein said peroxidase is at least one of soybean peroxidase and horseradish root peroxidase. 
     
     
       12. The electrode strip of  claim 1  wherein said at least one enzyme is one of creatine amidinohydrolase, glucose oxidase and cholesterol oxidase. 
     
     
       13. The electrode strip of  claim 12  wherein said second reagent further includes a second enzyme when said at least one enzyme is one of creatine amidinohydrolase and cholesterol oxidase. 
     
     
       14. The electrode strip of  claim 13  wherein said second enzyme is sarcosine oxidase when said at least one enzyme is creatine amidinohydrolase. 
     
     
       15. The electrode strip of  claim 13  wherein said second enzyme is cholesterol esterase when said at least one enzyme is cholesterol oxidase. 
     
     
       16. The electrode strip of  claim 1  wherein said redox mediator is an inorganic or organic redox species. 
     
     
       17. The electrode strip of  claim 16  wherein said redox species is at least one of Fe(CN) 6   3− , Fe(CN) 6   4− , Fe(0,10-phenanthroline) 3   2+ , Fe(2,2′-bipyridine) 3   2+ , Co(NH 3 ) 6   2+ , Co(1,10-phenanthroline) 3   2+ , Co(2,2′-bipyridine) 3   2+ , Os(2,2′-bipyridine) 2 Cl + , Os(1,10-phenanthroline) 2 Cl + , Ru(2,2′-bipyridine) 2   2+ , Rh(2,2′-bipyridine) 2   2+ , cobalt phthalocyanine, ferrocenes, methylene blue, methylene green, 7,7,8,8-tetracyanoquinodimethane, tetrathiafulvalene, toluidine blue, meldola blue, N-methylphenazine methosulfate, phenyldiamines, 3,3′,5,5′-tetramethylbenzidine, pyrogallol, and benzoquinone. 
     
     
       18. The electrode strip of  claim 17  wherein said redox mediator is potassium ferrocyanide. 
     
     
       19. The electrode strip of  claim 1  wherein said enclosed channel is hydrophilic. 
     
     
       20. The electrode strip of  claim 1  wherein said enclosed channel has a volume of about 1.5 microliters. 
     
     
       21. The electrode strip of  claim 1  wherein said cover has a hydrophilic coating on at least one side. 
     
     
       22. The electrode strip of  claim 1  wherein said first reagent and said second reagent are made from a mixture having starting components in water comprising about 2 wt % of potassium ferrocyanide, about 1 wt % of methyl cellulose, about 0.02 wt % of said 1-octylphenoxypolyethoxyethanol, about 0.5 wt % of glucose oxidase, and about 0.5 wt % of soybean peroxidase. 
     
     
       23. The electrode strip of  claim 22  wherein said first reagent and said second reagent further includes about 0.1 wt % of an antioxidant. 
     
     
       24. The electrode strip of  claim 1  wherein said first reagent and said second reagent are made from a mixture having starting components in water comprising about 5 wt % of potassium ferrocyanide, about 1 wt % of methyl cellulose, about 0.02 wt % of t-octylphenoxypolyethoxyethanol, about 2 wt % of cholesterol oxidase, about 1 wt % of cholesterol esterase, and about 0.5 wt % of soybean peroxidase. 
     
     
       25. The electrode strip of  claim 24  wherein said first reagent and said second reagent further includes about 0.1 wt % of an antioxidant. 
     
     
       26. A device for measuring creatinine comprising:
   a creatine measuring electrode having a creatine reagent comprising two enzymes capable of catalyzing a reaction involving a substrate for the enzyme creatine, a redox mediator and a peroxidase enzyme capable of catalyzing a reaction involving said redox mediator wherein said creatine reagent is water soluble;        a creatinine measuring electrode having a creatinine reagent comprising three enzymes capable of catalyzing a reaction involving a substrate for the enzyme creatinine, a redox mediator and a peroxidase enzyme capable of catalyzing a reaction involving said redox mediator wherein said creatinine reagent is water soluble; and        a reference electrode.     
     
     
       27. The device of  claim 26  wherein said two enzymes are creatine amidinohydrolase and sarcosine oxidase. 
     
     
       28. The device of  claim 26  wherein said three enzymes are creatinine amidohydrolase, creatine amidinohydrolase and sarcosine oxidase. 
     
     
       29. The device of  claim 26  further comprising a sample chamber containing said creatine measuring electrode, said creatinine measuring electrode and said reference electrode. 
     
     
       30. A creatinine sensor comprising:
   a laminated body having a fluid sampling end and an electrical contact end;        a sampling end aperture at said fluid sampling end;        a sample fluid channel in communication between said fluid sample inlet and a vent opening, said sample fluid channel being adapted to collect a fluid sample through said sampling end aperture;        a creatine measuring electrode having a creatine reagent comprising two enzymes capable of catalyzing a reaction involving a substrate for the enzyme creatine, a redox mediator and a peroxidase enzyme capable of catalyzing a reaction involving said redox mediator wherein said creatine reagent is water - soluble;        a creatine - creatinine measuring electrode having a creatinine reagent comprising three enzymes capable of catalyzing a reaction involving a substrate for the enzyme creatinine, a redox mediator and a peroxidase enzyme capable of catalyzing a reaction involving said redox mediator wherein said creatinine reagent is water - soluble; and        a reference electrode wherein said creatine measuring electrode, said creatine - creatinine measuring electrode and said reference electrode are disposed within said sample fluid channel.     
     
     
       31. The creatinine sensor of  claim 30  wherein said two enzymes are creatine amidinohydrolase and sarcosine oxidase. 
     
     
       32. The creatinine sensor of  claim 30  wherein said three enzymes are creatinine amidohydrolase, creatine amidinohydrolase and sarcosine oxidase.