There are different possible glucuronides that can be used as substrates for the β-glucuronidase, depending on the type of detection needed (histochemical, spectrophotometrical, fluorimetrical). The most common substrate for GUS histochemical staining is 5-bromo-4-chloro-3-indolyl glucuronide (X-Gluc). X-Gluc is hydrolyzed by GUS into the product 5,5-dibromo-4,4-dichloro-indigo (diX-indigo). DiX-indigo will appear blue, and can be seen using light microscopy. This process is analogous to hydrolysis of X-gal by Beta-galactosidase to produce blue cells as is commonly practiced in bacterial reporter gene assays.
For other types of detection, common substrates are [https://pubchem.ncbi.nlm.nih.gov/compound/p-nitrophenyl-beta-D-glucuronide p-nitrophenyl β-D-glucuronide] for the spectrophotometric assay and [https://pubchem.ncbi.nlm.nih.gov/compound/4-Methylumbelliferyl-glucuronide 4-methylumbelliferyl-beta-D-glucuronide] (MUG) for the fluorimetric assay.