Test strip for determining concentration of creatinine in a body fluid sample

Abstract

Apparatus for determining concentration of creatinine in whole blood or plasma using a 1-methylhydantoinase (NMHase) catalyzed reaction, wherein the invention uses commercially available NMHase and thus eliminates prior art stabilization procedures for NMHase. Conveniently, known Trinder reagents and oxidative couplers are used for the indicator system. It has been found that by judiciously selecting the reagents used for the indicator system and/or varying the amount of enzyme NMHase that is loaded into the assay, the effect of the blank reaction can be minimized in the dynamic range of interest such that concentration of creatinine in normal and pathological levels can be measured directly without having to adjust for a blank reaction caused by bound N-methylhydantoin (NMH).

1 . A test strip for determining concentration of creatinine in a body fluid sample, comprising a reagent layer impregnated with 1-methylhydantoinase (“NMHase”) having 1-methylhydantoin (“NMH”) bound thereto, whereby, when wetted with the sample, the test strip produces a response substantially proportional to the concentration of creatinine in the sample without being substantially affected by a blank reaction.  
   
   
       2 . The test strip of  claim 1 , wherein said reagent layer further comprises an oxidative coupling agent, said test strip further comprising an auxiliary reagent layer in fluid communication with said reagent layer, said auxiliary layer having impregnated therein a chromogen which corresponds to said oxidative coupling agent.  
   
   
       3 . The test strip of  claim 2 , wherein said reagent layer is impregnated with a first aqueous solution having an alkaline pH and said auxiliary reagent layer is impregnated with a second aqueous solution having an acidic pH.  
   
   
       4 . The test strip of  claim 3 , wherein said alkaline pH is at least 2 pH units greater than said acidic pH.  
   
   
       5 . The test strip of  claim 2 , further comprising a third layer disposed between said reagent layer and said auxiliary reagent layer, said third layer providing fluid communication between said auxiliary layer and said reagent layer when wetted with the sample.  
   
   
       6 . The test strip of  claim 2 , wherein said auxiliary layer comprises MgCl 2 .  
   
   
       7 . The test strip of  claim 2 , wherein said auxiliary layer comprises a nucleoside triphosphate.  
   
   
       8 - 20 . (canceled)