CHEMBL5738704

DPPIV Enzymatic Activity Assay: DPPIV enzymatic activity assay. To assay baseline dipeptidyl peptidase-4 (DPPIV) activity, 40 ng of recombinant human DPPIV (rhDPPIV) (R&S system, #1180-SE) or 40 ng of recombinant mouse DPPIV (rmDPPIV) (R&S system, #954-SE) was incubated with 400 μM of H-Gly-Pro-pNA substrate (BACHEM, #L-1880) in a DPPIV assay buffer (25 mM Tris, pH 8.3) for 30 min at 37° C. protected from the light in 96-well black plates (Nunc, #237108). To assay DPPIV inhibition by test compounds, test compounds were pre-incubated with the enzyme for 15 min at 37° C. before starting the reaction by substrate addition in 96-well black plates (Nunc, #237108). Para-nitroaniline (pNA) release was detected by measuring absorbance at 405 nm using a Multifunction Microplate Reader (Synergy 4, Biotek). All measurements were carried out in triplicate. Val-boroPro, a non-specific prolyl peptidase inhibitor, was used as a positive control.