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DPP-4 enzyme assays The enzyme assay was performed in flat-bottom 96- well microplates, where each reaction consisted of 100 L of either 12.5% human, platypus or echidna serum, 0.5 mM of H-Ala-Pro-p-nitroanilide (Bachem) as a substrate in the presence/absence of 100 M of DPP-4 inhibitor P32/98 (Santa Cruz Biotech) in 100 mM sodium phosphate buffer (PH 7.4) supplemented with 150 mM NaCl and 0.2% Tween-20 with a total volume of 200 M

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Total length 15 hours 40 minutes

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