| Solution Information | help | |
| Enzyme: | Cannabinoid receptor 1 | |
| inhibitor: | BDBM76072 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Materials: 1) 384-well plates, black with clear bottom (Greiner# 781091) 2) U2OS (Human Osteosarcoma) cell line stably expressing the Beta-arrestin GFP and the CB1 receptor 3) Culture Media: MEM with L-glutamine, Pen-strep, 10% Fetal Bovine Serum and selection antibiotics - 200ug/ml G418 and 50ug/ml Zeocin 4) Positive Control Working Solution: WIN55212 (Sigma W102 25mg) - 5mM stock in DMSO). 5) Negative Control Working Solution: AM251 (Sigma A6226-10mg) 5mM stock in DMSO 6) Test compounds from dry powders, working solution: 10mM in 100% DMSO 7) Fixative Working Solution: 6% Paraformaldehyde (PFA) diluted in PBS. 8) Nuclear Stain Working Solution: DAPI (Invitrogen, D1306) diluted to 150ng/ml in DAPI buffer (10mM TRIS, 10mM EDTA, 100mM NaCl, pH 7.4). Assay Procedure: 1) 45ul of cell suspension (200,000 cells/ml in culture medium) was dispensed in each well of the assay plates using a Wellmate bulk dispenser. 2) Plates are incubated overnight or approximately 20 hours at 37 deg | |
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