| Assay Method Information | |
| | Test of Inhibitory Activity on SGLT1 |
| Description: | Test Purpose:the following methods can be used to determine the inhibitory activity of the compounds described in the invention on SGLT-1.Test Materials:14C-AMG solution was purchased from PerkinElmer, Cat. No. NEZ080001MC;α-Methylglucoside was purchased from Sigma, Cat. No. M9376-100G.N-methyl-D-glucosamine was purchased from Sigma, Cat. No. M2004-100G.Phloridzin was purchased from Sigma, Cat. No. P3449-1G.96-Well cell culture plate was purchased from Corning, Cat. No. 3903.Test Method:mock-transfected FIP-in CHO cells (3×104) and CHO cells expressing human SGLT1 gene were seeded into 96-well plates respectively. The cells were incubated for 12 hours. Each well of the 96-well plate was washed with 150 μL of sodium-free buffer once. To each well was added 50 μL of sodium-containing buffer containing test compounds of different concentrations and 0.5 μM [14]-AMG. The incubation mixture was incubated at 37° C. for 1 hour. To each well was added 150 μL of precooled sodium-free buffer to terminate the reaction. The cell pellet was washed with sodium-free buffer three times and the residual liquid in well was removed. To each well was added 20 μL of precooled 100 mM NaOH. The 96-well plate was vibrated at 900 rpm for 5 minutes. Scintillation fluid (80 μL) was added to each well which was then vibrated at 600 rpm for 5 minutes. The amount of 14C-AMG was quantitatively detected using liquid scintillation. |
| Affinity data for this assay | |
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