In our efforts to improve the identification of phosphopeptides by MS we have used peptide IEF on IPG strips. Phosphopeptides derived from trypsin digests of single proteins as well as complex cellular protein mixtures can be enriched by IEF and recovered in excellent yields at the acidic end of an IPG strip. IPG peptide fractionation in combination with MS/MS analysis has allowed us to identify phosphopeptides from tryptic digests of a cellular protein extract.