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The clarified supernatant was filtered through a 0.4 µM filter to remove the insoluble fraction and supplemented with 30 mM Imidazole pH 7.0 immediately before loading at a flow rate of 2 mL/min onto a 5 mL Ni-NTA immobilized metal affinity chromatography (IMAC) column (Bio-Rad) equilibrated in Buffer A (50 mM Tris-Cl, 300 mM NaCl, 0.01% LMNG/0.001% CHS, pH 7.0)

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When levels are not optimal, it may influence stamina, concentration, and overall vitality