| Solution Information | help | |
| Enzyme: | G-protein coupled receptor 35 | |
| inhibitor: | BDBM65728 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay materials: 1) HEK-293-T NFKB-Luc cell line obtained from the assay provider's laboratory. 2) g-tri-DAP (Ana Spec cat #60774) obtained from assay provider's laboratory. 3) SteadyGlo (Promega) Primary Screen and Single-concentration confirmation Day 1 Procedure 1) Harvest HEK-293-T NFKB-Luc at 100% confluency 2) Dispense 3 uL (6000 cells)/well to every well of a 1536 TC-treated white plate (Corning # 3727). 3) Spin down plates at 1000 rpm for 1 min in an Eppendorf 5810 centrifuge. 4) Using a HighRes biosolution pintool equipped with V&P Scientific pins, stamp 10nl of 2mM cmpds in DMSO (col 5-48) and 10nl DMSO controls (col 1-4) to plates 5) Lid Plates. Incubate cells for 1 hour at room temp. 6) Dispense 2 uL/well of g-tri-DAP (1.875 ug/mL) in assay media containing 1.375% DMSO to columns 3-48. 7) Spin down plates for 30 sec in an Eppendorf 5810 centrifuge. 8) Lid Plates. Incubate overnight (16 hours) in 37 oC 5% CO2 incubator Day 2 Procedure 1) Equibrate plates to | |
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