Waters SILAC Hi3 Standards - Manual (Page 3)

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[ CARE AND USE MANUAL ]
Figure 2. Base Peak Intensity Chromatograms: 1D MS E – 25 fmol Hi3 SILAC PhosB.
CV 6.57% CV 6.57%
L a be l P e ptide S e qu e n c e [ M + H ] +
1 TC
am
AYTNHTVLPEALER 1884.9167
2 VFADYEEYVK 1270.6094
3 LITAIGDVVNHDPVVGDR 1900.0193
4 LYPNDNFFGK 1450.7104
5 LLSYVDDEAFIR 1470.7469
L a be l P e ptide S e qu e n c e [ M + H ] +
1 1 TC
am
AYT YY NHTVLPEALER TC
am
AYTNHTVLPEALER 1884.9167 1884.9167
2 2 VFADYEEYVK VFADYEEYVK 1270.6094 1270.6094
3 3 LITAIGDVVNHDPVVGDR LITAIGDVVNHDPVVGDR 1900.0193 1900.0193
4 4 LY L PNDNFFGK LYPNDNFFGK 1450.7104 1450.7104
5 5 LLSYVDDEAFIR LLSYVDDEAFIR 1470.7469 1470.7469
1 1
4 4
3 3
2 2
5 5
SILAC Hi3 Standards
L a be l P e ptide S e qu e n c e [ M + H ] +
1 NNPVLIGEPGVGK 1300.723
2 LPQVEGTGGDVQPSQDVLR 2004.016
3 VIGQNEADAVSNAIR 1664.873
4 VTDAEIAEVLAR 1295.696
5 AIDLIDEAASSIR 1382.727
6 AIQQQIENPLAQQILSGELVPGK 2481.369
L a be l P e ptide S e qu e n c e [ M + H ] +
1 1 NNPVLIGEPGVGK NNPVLIGEPGVGK 1300.723 1300.723
2 2 LPQVEGTGGDVQPSQDVLR LPQVEGTGGDVQPSQDVLR 2004.016 2004.016
3 3 VIGQNEADAVSNAIR VIGQNEADAVSNAIR 1664.873 1664.873
4 4 VTDAEIAEVLAR VTDAEIAEVLAR 1295.696 1295.696
5 5 AIDLIDEAASSIR AIDLIDEAASSIR 1382.727 1382.727
6 6 AIQQQIENPLAQQILSGELVPGK AIQQQIENPLAQQILSGELVPGK 2481.369 2481.369
1 1
2 2
3 3
4 4
5 5
6 6
Figure 1. Base Peak Intensity Chromatograms: 1D MS E – 25 fmol Hi3 SILAC ClpB E. coli.
The following is an example of the standards run on a
Waters SYNAPT G2-S HDMS mass spectrometer and the
corresponding results.
Conditions:
Solvent A:
0.1% formic acid in water
Solvent B: 0.1% formic acid in acetonitrile
Weak wash: 3% ACN w/ 0.1% TFA
Trapping: 10 µL/min for 3 min, 10 µL loop
Trap: ACQUITY UPLC ® PST C
18
nanoACQUITY ® Trap,
5 μm, 180 μm x 20 mm
Gradient Profile:
Time
Flow Rate
(µL/min)
%A %B Curve
Initial 0.500 95.0 5.0 6
54.00 0.500 60.0 40.0 6
55.80 0.500 15.0 85.0 6
59.40 0.500 15.0 85.0 6
61.20 0.500 95.0 5.0 6
72.00 0.500 95.0 5.0 6
Table 3. Base Peak Intensity Chromatograms: 1D MS E – 25 fmol
SILAC Hi3 ClpB E. coli.
IV. USING T HE SILAC HI3 DIGESTION STANDARDS FOR QUANTIFICATION