Waters SILAC Hi3 Standards - Manual (Page 2)

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[ CARE AND USE MANUAL ]
II. STORAGE AND STABILIT Y
Lyophilized peptides generally have excellent stabilities, often
showing little or no degradation after a few years at -20 °C. Long
term storage (>1 year) should be at -80 °C desiccated, medium
term storage (1–12 months) should be at -20 °C desiccated, short
term storage (<1 month) may be at 4 °C. Once reconstituted the
solution should be used immediately to avoid degradation of
peptides which would compromise the quantitative benefit.
SILAC Hi3 Standards
III. RECONSTITUTION OF T HE
SILAC HI3 DIGESTION STANDARDS
The SILAC Hi3 Standards are originally in a concentration of
1 nmol/vial of either the ClpB_ECOLI or the PYGM_RABIT.
The lyophilized peptides were re-suspended in 1000 µL of
3% acetonitrile w/ 0.1% Trifluoroacetic acid. The resulting
concentration was 1 pmol/µL. The SILAC Hi3 standards were then
diluted to 25 fmol/µL, i.e., 25 µL of each solution was diluted to
1000 µL. Because peptides can adhere to glass and other surfaces
resulting in significant losses from highly dilute solutions,
further dilutions should be made just prior to addition to samples
and excess diluted standard should be discarded. Unused stock
solutions may be stored in a freezer for a brief time. Analysis of
small amounts of standard (ca. 50 fmol) should only be carried
out in the presence of a sample matrix such as E. coli digest of a
serum digest (60 ng/µL or greater).
Table 1. SILAC Hi3 Phos B Standard
Peptide Sequence Average Molecular Weight Concentration
SILAC Phos B Sequence 1 H2N-VLYPNDNFFEGK-OH 1449.7066 1 nmol/vial
SILAC Phos B Sequence 2 H2N-TC*AYTNHTVLPEALER-OH 1883.9227 1 nmol/vial
SILAC Phos B Sequence 3 H2N-IGEEYISDLDQLRK-OH 1685.8762 1 nmol/vial
SILAC Phos B Sequence 4 H2B-LLSYVDDEAFIR-OH 1449.7531 1 nmol/vial
SILAC Phos B Sequence 5 H2N-LITAIGDVVNHDPVVGDR-OH 1899.0242 1 nmol/vial
SILAC Phos B Sequence 1 H2N-VLYPNDNFFEGK-OH 1269.6055 1 nmol/vial
The cysteine in sequence 2 is carbamidomethylated.
Table 2. SILAC Hi3 E. coli Standard
Peptide Sequence Average Molecular Weight Concentration
SILAC E. coli ClpB Sequence 1 H2N-VIGQNEAVDAVSNAIR-OH 1664.8873 1 nmol/vial
SILAC E. coli ClpB Sequence 2 H2N-NNPVLIGEPGVGK-OH 1300.7276 1 nmol/vial
SILAC E. coli ClpB Sequence 3 H2N-AIDLIDEAASSIR-OH 1382.7433 1 nmol/vial
SILAC E. coli ClpB Sequence 4 H2N-VTDAEIAEVLAR-OH 1295.7113 1 nmol/vial
SILAC E. coli ClpB Sequence 5 H2N-AIQQQIENPLAQQILSGELVPGK-OH 2482.3751 1 nmol/vial
SILAC E. coli ClpB Sequence 1 H2N-LPQVEGTGGDVQPSQDLVR-OH 2004.0304 1 nmol/vial