A substrate formulation which produces chemiluminescence in the presence of a phosphatase enzyme which comprises in an aqueous solution: a) 0.01 mM-50 mM of a chemiluminescent compound of formula I or a salt thereof: wherein A is C 1-6 haloalkyl, naphthyl, phenyl, substituted phenyl, or heteroaryl, wherein substituted phenyl comprises from 1 to 3 halo, C 1-6 alkyl, C 1-6 alkoxy, C 1-6 haloalkyl, C R 15, CN or NO 2 substituents; R 1 is selected from the group consisting of C 5-14 aryl, C 1-6 alkyl, C 1-6 haloalkyl, and C 5-14 aralkyl groups; R 7 -R 14 are independently H, C 1-6 alkoxy, halo, C 1-4 alkyl, or R 7 -R 8 or R 8 -R 9 or R 9 -R 10 or R 11 -R 12 or R 12 -R 13 or R 13 -R 14, can be joined together as a carbocyclic or heterocyclic ring system comprising at least one 5 or 6-membered ring; R 15 is C 1-6 alkyl; each M is independently selected from H, or an alkali metal, alkaline earth metal, transition metal, ammonium, phosphonium, organic amine salt, amino acid salt or; Z is O or S; and n is 0, 1, or 2; b) 0.01-200 uM of a cationic aromatic compound; c) 1 uM -10 mM of a background reducing agent; and d) 0.05 - 20 g/L of an ether-linked nonionic surfactant that does not contain a sulfate, sulfonate, phosphate or carboxylate ester group; or a hydrophilic polymer, wherein the ether-linked nonionic surfactant is of formula: wherein R is selected from C 6-22 alkyl, cycloalkyl, C 6-22 alkyl-substituted cycloalkyl, and mono- or di- C 6-22 alkyl-substituted phenyl; n is a number from 2-200; X is selected from O or S; and Y is selected from H or C 1-4 alkyl. a) 0.01 mM-50 mM of a chemiluminescent compound of formula I or a salt thereof: wherein A is C 1-6 haloalkyl, naphthyl, phenyl, substituted phenyl, or heteroaryl, wherein substituted phenyl comprises from 1 to 3 halo, C 1-6 alkyl, C 1-6 alkoxy, C 1-6 haloalkyl, C R 15, CN or NO 2 substituents; R 1 is selected from the group consisting of C 5-14 aryl, C 1-6 alkyl, C 1-6 haloalkyl, and C 5-14 aralkyl groups; R 7 -R 14 are independently H, C 1-6 alkoxy, halo, C 1-4 alkyl, or R 7 -R 8 or R 8 -R 9 or R 9 -R 10 or R 11 -R 12 or R 12 -R 13 or R 13 -R 14, can be joined together as a carbocyclic or heterocyclic ring system comprising at least one 5 or 6-membered ring; R 15 is C 1-6 alkyl; each M is independently selected from H, or an alkali metal, alkaline earth metal, transition metal, ammonium, phosphonium, organic amine salt, amino acid salt or; Z is O or S; and n is 0, 1, or 2; A is C 1-6 haloalkyl, naphthyl, phenyl, substituted phenyl, or heteroaryl, wherein substituted phenyl comprises from 1 to 3 halo, C 1-6 alkyl, C 1-6 alkoxy, C 1-6 haloalkyl, C R 15, CN or NO 2 substituents; R 1 is selected from the group consisting of C 5-14 aryl, C 1-6 alkyl, C 1-6 haloalkyl, and C 5-14 aralkyl groups; R 7 -R 14 are independently H, C 1-6 alkoxy, halo, C 1-4 alkyl, or R 7 -R 8 or R 8 -R 9 or R 9 -R 10 or R 11 -R 12 or R 12 -R 13 or R 13 -R 14, can be joined together as a carbocyclic or heterocyclic ring system comprising at least one 5 or 6-membered ring; R 15 is C 1-6 alkyl; each M is independently selected from H, or an alkali metal, alkaline earth metal, transition metal, ammonium, phosphonium, organic amine salt, amino acid salt or; Z is O or S; and n is 0, 1, or 2; b) 0.01-200 uM of a cationic aromatic compound; c) 1 uM -10 mM of a background reducing agent; and d) 0.05 - 20 g/L of an ether-linked nonionic surfactant that does not contain a sulfate, sulfonate, phosphate or carboxylate ester group; or a hydrophilic polymer, wherein the ether-linked nonionic surfactant is of formula: wherein R is selected from C 6-22 alkyl, cycloalkyl, C 6-22 alkyl-substituted cycloalkyl, and mono- or di- C 6-22 alkyl-substituted phenyl; n is a number from 2-200; X is selected from O or S; and Y is selected from H or C 1-4 alkyl.