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4 protocols using imp 1088

1

Cell growth assay with 2-hydroxymyristate and IMP-1088

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Cells were grown in high-glucose DMEM supplemented with 10% fetal bovine serum, 2 mM glutamine, 5 mM non-essential amino acids and 1% (v/v) MEM vitamins (Thermo Fisher Scientific). Growth measurements were performed in 96-well plates and quantified by staining with the sulforhodamine B (SRB) dye, as previously described [3 (link)]. Absorbance was measured at 560 nm with a microplate reader. As indicated in the legend of figures, cells were grown in the presence of increasing concentrations of 2-hydroxymyristate (Cayman, Ann Arbor, MI, USA) and IMP-1088 (Cayman).
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2

Myristoylation Detection and Inhibition

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Myristoylation was detected using a myristoylated protein assay kit (abcam, USA) following the manufacturer’s instructions. Cells without myristic acid labeling served as background controls. To test the myristoylation inhibitor, cells were pre-incubated with 1 µM n-myristoyltransferase inhibitor IMP-1088 (Cayman Chemical, USA) for 1h before stimulation with 100 ng/ml upLPS overnight.
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3

Iodixanol Density Gradient Cell Fractionation

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IMP-1088 was purchased from Cayman Chemical (Cat# 25366–1). Iodixanol (Opti-Prep Density Gradient Medium) was purchased from Millipore-Sigma (Cat# D1556). SMARTPool ON-TARGETplus and non-targeting control #2 siRNAs were purchased from Dharmacon.
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4

Ferroptosis Regulators Protocol

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Lip-1 (Selleckchem, cat. no. S7699), doxycycline (Dox; Sigma, cat. no. D9891), RSL3 (Cayman, cat. no. 19288), BSO (Sigma, cat. no. B2515), iFSP1 (ChemDiv, cat. no. 8009-2626), icFSP1 (ChemDiv, cat. no. L892-0224 or custom synthesis by Intonation Research Laboratories), erastin (Merck, cat. no. 329600), ML210 (Cayman, cat. no. Cay23282-1), FIN56 (Cayman, cat. no. Cay25180-5), FINO2 (Cayman, cat. no. Cay25096-1), deferoxamine mesylate salt (DFO; Sigma, cat. no. 138-14-7), ferrostatin-1 (Fer-1; Sigma, cat. no. SML0583), zVAD-FMK (zVAD; Enzo Life Sciences, cat. no. ALX-260-02), necrostatin-1s (Nec-1s; Enzo Life Sciences, cat. no. BV-2263-5), MCC950 (Sigma, cat. no. 5381200001), olaparib (Selleckchem, cat. no. S1060), staurosporine (STS; Cayman, cat. no. 81590), recombinant human tumour necrosis factor (TNF; R&D Systems, cat. no. NBP2-35076), Smac mimic (BV-6; Selleckchem, cat. no. S7597), nigericin (Thermo Fisher, cat. no. N1495), IMP-1088 (Cayman, cat. no. Cay25366-1) and lipopolysaccharide (LPS; Sigma, cat. no. L2880) were used in this study.
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