| Assay Method Information | |
| | In Vitro Renin IC50 Determinations (Tandem GFP FRET Assay) |
| Description: | The renin assay utilized a tandem Green Flourescent Protein (tGFP) substrate that was hydrolyzed by human rennin. The tGFP substrate contained a nine amino acid (Ile-His-Pro-Phe-His-Leu-Val-Ile-His) recognition sequence for human renin flanked by two GPF proteins (W1B and Topaz). Human rennin cleaves the leucine-valine site of the substrate linker. Once cleaved, the emission ratio changes. The change was monitored by the ratio of 530 nm (topaz) over 475 nm (W1B) with the excitation set at 432 nm and the cutoff at 515 nm. The assay used a 384 well plate format that was read using a Gemini XS Fluorometric plate reader (Molecular Devices). Compounds were screened at a starting concentration of 10 uM and used a fourfold eleven-point dilution regiment. |
| Affinity data for this assay | |
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