Serum (stored at −80°C in 100 µL aliquots), was thawed in a 25°C water bath for 10 minutes, then stored on ice prior to sample dilution. Samples were mixed by gentle vortexing for 8 seconds. A 6% serum sample solution was prepared by dilution into 0.94× SB17 supplemented with 0.6 mM MgCl2, 1 mM trisodium EGTA, 0.8 mM AEBSF, and 2 µM Z-Block. A portion of the 6% serum stock solution was diluted 10-fold in SB17 to create a 0.6% serum stock. 6% and 0.6% stocks are used to detect high- and low-abundance analytes, respectively.
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