| Assay Method Information | |
| | Inhibitory Activity on Proton Pump (H+/K+-ATPase) Activity |
| Description: | 60 ul of enzyme reaction solution (a test compound at a given concentration, 3 g of gastric mucosal microsomes, 3 mM of MgSO4, 5 mM of KCl, 40 mM of Tris-HCl pH 6.4, and 0.5 mM of adenosine triphosphate) was added to each well of a 96-well plate and reacted at 37° C. for 60 minutes, and 30 μl of 10% SIDS was added to stop the enzyme reaction. 200 ul of detection solution (in which 10% L-ascorbic acid with pH 5.0 was mixed with 35 mM of ammonium molybdate tetrahydrate/15 mM of zinc acetate with pH 5.0 at the ratio of 4:1) was added and reacted at 37 C. for 30 minutes, and the absorbance value was measured at 750 nm by using a microplate reader (MULTISKAN GO, Thermo). The inhibition rate of proton pump activity was expressed as I050, which is 50% inhibition of enzyme activity, by using GraphPad Prism 5. |
| Affinity data for this assay | |
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