Identifying peptides based on HPLC retention

[pipettemonkey]

Quite a bit of information is available for predicting peptide retention (C18 TFA/MeCN) based on amino acid side chains and their contribution to hydrophobicity. However, knowledge of C18 retention alone is insufficient for identifying the peptide from a database.

What other modes, when combined with C18 retention behavior, would reduce the number of possibilities (most dramatically) when screening peptide databases?

[Uwe Neue]

A well-established data base on the packing that you are using could be helpful, but the overall better tool is MS.

[Kostas Petritis]

It will mostly depend on the size of your database... Are we talking about a protein digest or a global proteome of an organism? You can play with reverse phase time prediction as well as pI prediction (if you can use isoelectric focusing as an additional dimension of separation) but those won't give you the necessary confidence if your database is even a mixture of proteins...

When you add the MS information, then you can start being much more confident...