| Solution Information | help | |
| Enzyme: | Integrin alpha-4 | |
| inhibitor: | BDBM40201 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Binding assays are performed in a format adapted for multiwell plates. 384-well microplates are configured with control wells (control cells with no compound in columns 1, 2, 12, and 13 then buffer only in columns 14, 15). Per row, a nine serial dilution of compounds are added to columns 3 to 11. The fluorescent ligand utilized to evaluate affinity state of VLA-4 is LDV-FITC containing small molecule, 4-((N'-2-methylphenyl)ureido)-phenylacetyl-L-leucyl-L-aspartyl-L-valyl-L-prolyl-L-alanyl-L-alanyl-L-lysine-FITC. This dose-response plate format is repeated for three types of plates: 1) No compound (cells with compound carrier, DMSO), 2) Cells with 1 nM LDV-FITC, 3) Cells with 1 nM LDV-FITC plus 0.5 mM Mn2+. Additions to wells are in sequence as follows. Dilutions of compounds are made in 384-well plates with HHB (30 mM Hepes, 110 mM NaCl, 10 mM KCl, 10 mM Glucose, 1 mM Mg2+, 1.5 mM Ca2+) plus 0.1% Bovine Serum Albumin (BSA) at 2x the final concentration. Each subsequent compou | |
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