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Solution Informationhelp
Enzyme: G-protein coupled receptor 35
inhibitor: BDBM75859
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 GPR35 receptor 3) Culture Media: MEM with L-glutamine, Pen-Strep, 10% Fetal Bovine Serum, 10mM Hepes, and selection antibiotics - 200ug/ml G418 and 50ug/ml Zeocin 4) DMSO Working Solution: 100% DMSO 5) Agonist Working Solution: Zaprinast (Alexis #ALX-430-020-M010,10mg) 5mM stock in DMSO diluted to 100uM in PBS. 6) Test compounds from dry powder 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 degree C and
 
 

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Last update November 1, 2007
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