| Assay Method Information | |
| | AlphaScreen Binding Assay |
| Description: | This assay can be used to examine the potency with which compounds inhibit the protein-protein interaction between SOS1 and KRAS G12D. This demonstrates the molecular mode of action of compounds. Low IC50 values are indicative of high potency of the SOS1 inhibitor compound in this assay setting:Reagents:GST-tagged SOS1 (564_1049_GST_TEV_ECO) produced in-houseGST-TEV-SOS1 (564-1049) is purchased from Viva Biotech Ltd.6 His-Tev-K-RasG12D(1-169)Avi is purchased from Xtal BioStructures, Inc. (Lot #X129-110)GDP (Sigma Cat No G7127)AlphaLISA Glutathione Acceptor Beads (PerkinElmer, Cat No AL109)AlphaScreen Streptavidin Donor Beads (PerkinElmer Cat No 6760002)Assay plates: Proxiplate-384 PLUS, white (PerkinElmer, Cat No 6008289)Assay Buffer:1 PBS0.1% BSA100 uM EDTA or without EDTA (IC50s in the tables are measured without EDTA unless they are marked with an asterisk)0.05% Tween 20KRAS SOS1 GDP Mix:10 nM (final assay concentration) KRAS G12D, 10 uM (final assay concentration) GDP and 5 nM (final assay concentration) GST-SOS1 are mixed in assay buffer prior to use and kept at room temperature. |
| Affinity data for this assay | |
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