Heparanase (HPSE) is an endo-β-glucuronidase involved in extracellular matrix remodeling in rapidly healing tissues, most cancers and inflammation, and viral infection. Its importance as a therapeutic target warrants further study, but such is hampered by a lack of research tools. To expand the toolkits for probing HPSE enzymatic activity, we report the design of a substrate scaffold for HPSE comprised of a disaccharide substrate appended with a linker, capable of carrying cargo until being cleaved by HPSE. Here exemplified as a fluorogenic, coumarin-based imaging probe, this scaffold can potentially expand the availability of HPSE-responsive imaging or drug delivery tools using a variety of imaging moieties or other cargo. We show that electronic tuning of the scaffold provides a robust response to HPSE while simplifying the structural requirements of the attached cargo. Molecular docking and modeling suggest a productive probe/HPSE binding mode. These results further support the hypothesis that the reactivity of these HPSE-responsive probes is predominantly influenced by the electron density of the aglycone. This universal HPSE-activatable scaffold will greatly facilitate future development of HPSE-responsive probes and drugs.