Hi,
Despite having worked in this area for a long time, I'm struggling with a few
finer points of phosphopeptide enrichment.
1. I thought our TiO2 and newer CaTiO3 methods were pretty fool-proof, but I
have some new people struggling to get high reproducibility.
Do people have experience with some of the pre-packaged, optimized systems,
like the Pierce High Select Fe-NTA columns?
How about magnetic bead systems vs spin-filters vs batch? Does anyone have a
clear preference either for ease of use or robustness and efficiency?
2. We have been working with some samples derived from fat tissue and are
getting horrible phosphopeptide enrichment. A positive control with equal
amount of HeLa peptides looks fine.
Anyone have any thoughts why this should be? I would have thought that any
lipids, including phospholipids would be lost to the C18 during pre-enrichment
clean up.
