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The supernatant was then loaded on top of a 12 mL 1050% sucrose gradient (25 mM HEPES pH 7.5, 5 mM MgCl 2 , 0.1 M KCl, 2 mM DTT, 0.1 mg/mL cycloheximide) and spun in an ultracentrifuge at 35,000 RPM at 4 C for 2 h
Higher concentrations (10mg/mL or greater) reduce injection volume but amplify measurement erroreach 0.01mL syringe increment represents 100mcg peptide, introducing 2040% relative error for sub-250mcg doses
Amino acid metabolism and signalling pathways: potential targets in the control of infection and immunity
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