Fig. 5.

N-glyco FASP applied to blood plasma. A, Blood plasma was mixed with the breast cancer cell secretome super-SILAC mix. The enrichment of N-glycosylated peptides, and thereby removal of nonglycosylated peptides, reduced the intensity of abundant plasma proteins by about 200 times. B, Distribution of ratios of N-glycosylation sites between the super-SILAC internal standard and plasma. Sites without a signal in the light or heavy SILAC channel were assigned a Log2 ratio of –7 and 9, respectively. Overlapping sites with the secretome data set are labeled in red, whereas classical plasma proteins (as taken from Anderson et al. (6)) are labeled in pink. C, D, E, example MS spectra of N-glycosylated peptides with differing ratios. C, IHVTIYN(de)CSFGR (3+), plexin B2, average ratio −1.75; D, FEAEHISN(de)YTAIIISR (3+), semaphorin 4B, average ratio 0.0; E, IDSTGN(de)VTNEIR (2+), attractin, average ratio 4.3.