Dry analytical element for acetaminophen assay

A dry multilayer analytical element for the determination of acetaminophen in an aqueous fluid, comprising a support having thereon, in order from said support and in fluid contact:

• (a) a layer having therein (i) an arylacylamidase enzyme;
• (ii) a ferricyanide capable of oxidatively coupling paraaminophenol to a color-forming coupling agent to form a color compound;
• and (iii) a water-soluble, color-forming coupling agent, wherein at least one layer comprises gelatin which does not undergo hardening due to ferricyanide-induced chemical crosslinking, and wherein said water-soluble, color-forming coupling agent has the general formula:
• wherein R is a water-solubilizing group selected from (1) —(CH2)nX, where n is 1 to 5, X is either (I) —SO3M where M is hydrogen, an alkali metal, an alkaline earth metal or an ammonium (NH4+) cation, or (II) (—OCH2CH2)yOH where y is 2 to 5;
• and (2) —N(R7)3+Z— where each R7 is independently selected from alkyl of 1 to 4 carbon atoms, and Z is an acid anion;
• R1 and R6 are taken together to represent an ethylene, trimethylene, or tetramethylene group which forms a partially saturated ring;
• and
• R2, R3, and R4 are independently selected from hydrogen, alkyl of 1 to 4 carbon atoms, and alkoxy of 1 to 4 carbon atoms;
• (b) a porous spreading layer;
• and
• (c) a buffer which maintains the pH of the element in a range of between about 6.5 to 8.5.