Measurement of total protein synthesis

1 µC of Perkin Elmer EasyTag ³⁵S labeled methionine (Product # NEG709A) was mixed with 5 µL 288 µmol unlabeled methionine and 24 µL media. At the time of capture, 900 µL of culture was added to methionine mix, and was labeled on a shaker for 1 min at 37°C. After labeling, 100 µL of ice-cold 50% trichloracetic acid (TCA) was added to the sample, which was vortexed and placed on ice for at least 20 min to allow precipitation. Samples were then counted by running 100 µL of sample through a 25 mm APFC glass fiber filter (Millipore APFC02500, Hainesport NJ) pre-wetted with 750 µL of 5% TCA on a vacuum stand, and washing three times with 750 µL 5% TCA and three times with 750 µL 100% ethanol. Filters were then placed in MP Ecolume scintillation fluid and counted.