Figure 2. Affect of acetonitrile in TiO₂ loading buffer.

One pmol of a tryptic digest of α- and β-casein was loaded onto TiO₂ using the MDLC similarly as in Fig. 1, except that different concentrations of acetonitrile (ACN) were used in the loading buffer and wash buffer, and peptides were eluted from TiO₂ using 200 mM NH₄HCO₃ pH 9. A comparison of 0-80 % acetonitrile (ACN) in both loading and wash buffers (all in the presence of 0.1% formic acid) is presented. Spectra count results of the four most prominent phosphopeptides, and the summary data of all identified phosphopeptides and the ratio of phosphopeptides to unmodified peptides are presented for each concentration of ACN used. Spectra count results are normalized to those observed for the 0% ACN buffer. Non-normalized data of all identified phosphopeptides is included in Supplemental Information (S-Table 8).