To: ABRF questions
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Dear ABRF's
I'm mainly involved in identification of 2D separated proteins.
The procedure for the identification that is used is in general the
following :
1. protein spot of intrest is digested in gel by trypsin
2. peptides are extracted
3. PorosR2 clean-up/concentration in a eppendorf Geloader tip
4. NanoSpray-ESI-Q-TOF- MS and MSMS, using micromass gold coated
needles
During NanoSpray-ESI-Q-TOF- MS the most important experiment is the MS
scan. Parent ion scanning can not be used (for the moment) on the Q-TOF
mass spec so that I'm totally dependent on the signal to noise in the
primary scan.
At this point a problem arise namely the peptide related multiple charged
ions are obscured by polymer like peaks especially in the region below
700m/z. These polymer peaks are probably extracted (spray solvent =
60%MeOH/1%HCOOH) from eppendorf tubes because working in glass tubes
gives a much cleaner spectrum. After a while (a few hours) even doubly
charged polymer peaks are observed in the MS spectrum
Here an overview of the most intense m/z values that are observed in the MS
scan :
serie 1 : 307.71 _ 409.17
serie 2 : 387.19_489.22_591.27_693.32_795.38
after 5 hours in eppendorf tube
serie 3 : singly charged series + doubly charge serie :
396.14_446.14_497.15_548.18_...
serie 4 : ........
serie 5 : ........
all these series differ from each other by 22 da!
accurate mass measurement of the polymer building block reveals a mass of
102.0513da!!
Has anyone observed the same problem or a similar problem?
thanks in advance,
Jos Raymackers
e-mail (innogenetics) : josray@innogenetics.be
e-mail (home) : josray@hotmail.com
INNOGENETICS NV
Industriepark 7 box 4
9052 Zwijnaarde Ghent
Belgium
tel. (office) : 0032-9-2410778
tel. (lab) : 0032-9-2410775
fax. : 0032-9-2410907