Dear colleagues,
I tried to analyze some phosphorylated peptides (around 8000 Da) in MALDI-TOF (positive linear mode).
However, it's difficult to obtain a high resoultion, very sharp peak. Instead, the peak base is usually broad with certain minor splits on top.
Would this be due to salts or is it an inherent property of the phosphoryl group?
Please shed some light on this. Thanks.
My first guess is that the sample prep (= purification, in this case) is not good enough.
The phospho group can give you trouble in the ionization process (if at all..)
But what you are talking about is resolution. First of all, 8kDa is more the size of a small protein than a peptide. You might consider the use of a different matrix (in case you were using HCCA).
For example, sinapic acid.
Or you might try the chlorinated version of HCCA, ClCCA. It is advertised for phosphopeptides.
Can you do delayed pulse extraction? if this is the case, what's the time there?
How did you prepare the sample?
Dear Kay Wong,
First thing 8kDa is comes under protein, 8000molecular weight means you should need to change instrumental data to enhance mode to avoid sensitive fragments and impurity, make sure your compound is pure, if not run HPLC and make it to pure, Then you first give M+ And M+H+ rest you hide by controlling instrumental instruction.. no you may see the peak what do you expect.
For this you do chlorinated version of ClCCA means you get better result.,
Then the peak is very broad its due to phosphoryl group effect doring ionistation because this group will stick and consume more time to response that time the peak getting broad and confusion peak was visible...
Hope no you clear....
Good Luck & Regards
R.Selva.................
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