Assay Method Information

Assay Name:  Fluorescent ACAT Assay
Description:  HepG2 cells were cultured in low-glucose DMEM supplement with 100 U/mL streptomycin and penicillin. Twenty thousand cells were plated per well on 96-well culture plates and grew overnight at 37 °C. Cells were incubated in medium containing either different samples, Sandoz 58-035 (positive control) or without sample (negative control and blank control) for 1 h. Then, NBD-cholesterol with final concentration of 2 µg/mL was added into the medium except for blank control and incubated overnight. NBDcholesterol was added from a 1 mg/mL stock in ethanol, and ethanol concentrations did not exceed 0.1%. After incubation, medium was removed, and the cells were washed three times with 20 µL PBS buffer. Plates were read fluorescence from the bottom using WALLAC Victor 2plate reader (PerkinElmer, Waltham, MA, USA) equipped with 485-nm excitation and 535-nm emission filters.
Affinity data for this assay
 

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