| Assay Method Information | |
| | Inhibition of ACSL5 Enzyme Activity Assay |
| Description: | Inhibition of ACSL5 enzyme activity was monitored with a coupled enzyme Amplex Red assay. The assay is carried out in 384-well microtiterplates. In a total volume of 40 L, compounds are preincubated with 20 nM ACSL5, in a buffer containing 150 mM MOPS-NaOH, 12 mM MgCl2, 0.03% (v/v) Triton X-100, 2.79 U mL-1 Peroxidase and 24 U mL-1 Acyl-CoA Oxidase (pH 7.6). The reaction is started by the addition of substrate and co factors (25 μM oleate, 30 μM CoA, 1 mM ATP and 50 μM Amplex Red) and the fluorescence detected after 10 minutes at 25° C. with a fluorescence reader (BMG-Fluostar; BMG-Technologies) using an excitation wavelength of 535 nm and an emission wavelength of 590 nm. |
| Affinity data for this assay | |
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