Tryptophanase-positive microorganisms (e.g. E. coli, S. faecalis, Proteus sp.) that cleave tryptophane hereby release indole, which may be detected by reaction with dimethylamino benzaldehyde by forming a dark red dye.

Microorganisms must be incubated in a glucose-free broth with high concentrations of tryptophane present. Kovacs’ Reagent is added to 24-48 h old cultures in a concentration of 0.2 to 0.5 ml per 5 ml broth. A colour shift to dark red shows presence of indole, thus confirming the inoculating colony as being indole-/tryptophanase-positive.