| Assay Method Information | |
| | Assay of Activity of Inhibiting KRAS (G12D) |
| Description: | 1) The concentration of a stock solution of the control compound was 1 mM, and the concentration of a stock solution of compounds to be assayed was 10 mM. 9 μL of the control compound and compounds to be assayed were transferred to a 384-LDV plate;2) The compounds on the LDV plate were serially diluted 3-fold with Bravo to 10 concentrations;3) 9 nL of the compounds on the LDV plate were transferred to an assay plate using ECHO;4) 3 μL of 3 nM Kras/0.5 nM TB-SA/30 nM BodipyGDP mixture and 3 μL of Ras buffer were sequentially added to each well of the assay plate using a Dragonfly automatic sampler, and the assay plate was centrifuged at 1000 rpm/min for 1 minute;5) The assay plate was incubated at room temperature for 1 hour;6) 3 μL of 120 nM SOS/9 mM GTP mixture was added to each well of the assay plate using a Dragonfly automatic sampler, and the assay plate was centrifuged at 1000 rpm/min for 1 minute;7) The assay plate was incubated at room temperature for 1 hour;8) The plate was read with Envision and data were recorded;9) The data were analysed using Excel and Xlfit, and IC50 of the compounds to be assayed were calculated. |
| Affinity data for this assay | |
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