| Assay Method Information | |
| | Inhibition Assay |
| Description: | Pretreatment buffer (140 mM choline chloride, 2 mM KCl, 1 mM CaCl2, 1 mM MgCl2, 10 mM HEPES/5 mM Tris, pH 7.4) was added to the stably expressing cells, followed by incubation for 20 minutes. The pretreatment buffer was removed and replaced by uptake buffer containing a test compound (1 mM methyl alpha-D-glucopyranoside (containing [14C]methyl alpha-D-glucopyranoside), 145 mM NaCl, 2 mM KCl, 1 mM CaCl2, 1 mM MgCl2, 10 mM HEPES/5 mM Tris, pH 7.4). Uptake reaction was performed at 37° C. for 30 minutes (SGLT1) or 60 minutes (SGLT2). After the reaction, the cells were washed twice with washing buffer (10 mM methyl alpha-D-glucopyranoside, 140 mM choline chloride, 2 mM KCl, 1 mM CaCl2, 1 mM MgCl2, 10 mM HEPES/5 mM Tris, pH 7.4), and then dissolved in a 0.25 M NaOH solution. A liquid scintillator (Perkin Elmer) was added and mixed well, followed by measurement of radioactivity using a beta-ray analyzer. For the control group, uptake buffer containing no test compound was prepared. |
| Affinity data for this assay | |
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