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The protein was finally eluted with buffer containing 20 mM HEPES, pH 7.5, 100 mM NaCl, 0.01% (w/v) MNG, 0.001% (w/v) CHS, 200 mg/ml Flag peptide, 5 mM EDTA and 1 M ghrelin or ibutamoren, and further purified by size-exclusion chromatography on a Superdex 200 Increase column (Cytiva) using buffer containing 20 mM HEPES pH 7.5, 100 mM NaCl, 0.01% (w/v) MNG, 0.001% (w/v) CHS, and 1 M ghrelin or ibutamoren

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