Mediator stabilized reagent compositions and methods for their use in electrochemical analyte detection assays

Abstract

Mediator stabilized reagent compositions and methods for their use in electrochemical analyte determination assays are provided. The subject reagent compositions include an enzyme, a redox mediator and a mediator-stabilizing buffer. Optionally, the reagent compositions may further include one or more of a wetting agent, detergent, enzyme cofactor and combinations thereof. Also provided are electrochemical test strips that include the subject reagent compositions, systems and kits that include the same as well as methods for using the same in analyte detection assays. The subject invention finds use in a variety of different applications, including glucose concentration determination applications.

1 - 38 . (canceled)  
     
     
         39 . A dry reagent composition comprising: 
 an enzyme;    a redox mediator;    an enzyme cofactor; and    a mediator-stabilizing buffer comprising a polycarboxylic acid having at least two carboxylic acid groups, present in an amount in the range of 0.01% wt/wt to 40% wt/wt sufficient to stabilize said redox mediator during storage of the dry reagent composition, wherein said mediator-stabilizing buffer has a low binding affinity for divalent metal ions, said low binding affinity for divalent metal ions being at least 2-fold less than a divalent metal ion binding affinity of an enzyme/cofactor complex formed from the enzyme and the cofactor of the dry reagent mixture.    
     
     
         40 . The reagent composition according to  claim 39 , wherein said polycarboxylic acid has from 2 to 10 carboxylic acid groups.  
     
     
         41 . The dry reagent composition according to  claim 39 , wherein said buffer has a pKa ranging from about 5 to about 8.  
     
     
         42 . The dry reagent composition according to  claim 41 , wherein said buffer has a pKa ranging from about 6 to about 7.  
     
     
         43 . The dry reagent composition according to  claim 39 , wherein said buffer is formulated to have a pH in the range from about 4 to 8.  
     
     
         44 . The dry reagent composition according to  claim 43 , wherein said buffer is formulated to have a pH in the range from about 6 to 7.  
     
     
         45 . The dry reagent composition according to  claim 39 , wherein at least two of said carboxylic groups are present in a cis formation.  
     
     
         46 . The dry reagent composition according to  claim 45 , wherein at least two of said carboxylic groups present in a cis formation are bonded to an ethylene moiety.  
     
     
         47 . The dry reagent composition according to  claim 46 , wherein said polycarboxylic acid is mellitic acid and salts thereof.  
     
     
         48 . The dry reagent composition according to  claim 39 , wherein said enzyme is an oxidizing enzyme.  
     
     
         49 . The dry reagent composition according to  claim 48 , wherein said oxidizing enzyme is chosen from an oxidase and a dehydrogenase.  
     
     
         50 . The dry reagent composition according to  claim 48 , wherein said enzyme is a dehydrogenase.  
     
     
         51 . The dry reagent composition according to  claim 48 , wherein said oxidizing enzyme is a glucose oxidizing enzyme.  
     
     
         52 . The dry reagent composition according to  claim 39 , wherein said mediator is ferricyanide.  
     
     
         53 . The dry reagent composition according to  claim 39 , wherein said reagent composition further comprises a stabilizer.  
     
     
         54 . The dry reagent composition according to  claim 53 , wherein said stabilizer is a carbohydrate.  
     
     
         55 . The dry reagent composition according to  claim 54 , wherein said carbohydrate is sucrose.  
     
     
         56 . An electrochemical cell comprising a dry reagent composition that includes: 
 an enzyme;    a redox mediator;    an enzyme cofactor; and    a mediator-stabilizing buffer comprising a polycarboxylic acid having at least two carboxylic acid groups, present in an amount in the range of 0.01% wt/wt to 40% wt/wt sufficient to stabilize said redox mediator during storage of the dry reagent composition, wherein said mediator-stabilizing buffer has a low binding affinity for divalent metal ions, said low binding affinity for divalent metal ions being at least 2-fold less than a divalent metal ion binding affinity of an enzyme/cofactor complex formed from the enzyme and the cofactor of the dry reagent mixture.    
     
     
         57 . The electrochemical cell according to  claim 56 , wherein said cell is present in an electrochemical test strip.  
     
     
         58 . A method for making an electrochemical test strip, the method comprising: 
 making an electrochemical strip that includes: 
 a dry reagent composition with: 
 an enzyme;  
 a redox mediator;  
 an enzyme cofactor; and  
 a mediator-stabilizing buffer comprising a polycarboxylic acid having at least two carboxylic acid groups, present in an amount in the range of 0.01% wt/wt to 40% wt/wt sufficient to stabilize said redox mediator during storage of the dry reagent composition, wherein said mediator-stabilizing buffer has a low binding affinity for divalent metal ions, said low binding affinity for divalent metal ions being at least 2-fold less than a divalent metal ion binding affinity of an enzyme/cofactor complex formed from the enzyme and the cofactor of the dry reagent mixture.  
 
   
     
     
         59 . A method for determining the concentration of an analyte in a sample, said method comprising: 
 (a) applying said sample to an electrochemical cell comprising a dry reagent composition that includes: 
 an enzyme;  
 a redox mediator;  
 an enzyme cofactor; and  
 a mediator-stabilizing buffer comprising a polycarboxylic acid having at least two carboxylic acid groups, present in an amount in the range of 0.01% wt/wt to 40% wt/wt sufficient to stabilize said redox mediator during storage of the dry reagent composition, wherein said mediator-stabilizing buffer has a low binding affinity for divalent metal ions, said low binding affinity for divalent metal ions being at least 2-fold less than a divalent metal ion binding affinity of an enzyme/cofactor complex formed from the enzyme and the cofactor of the dry reagent mixture;  
   (b) detecting an electrical signal produced by said cell; and    (c) relating said detected electrical signal to the concentration of said analyte in said sample.    
     
     
         60 . A method according to  claim 59 , wherein said analyte is glucose.  
     
     
         61 . A method according to  claim 59 , wherein said physiological sample is blood.