How many bands does one typically get in the second dimension of the ProteoSep fractionation?
A. This is a sample dependent answer, but for whole cell lysates you will see typically 1000-1500 “peaks” in the maps. This is essentially the same as that for silver staining. Each “peak” may contain more than one protein, however, and this leads to the possibility that after an additional LC/MS analysis the number of visible proteins could double or triple (or more) compared to gel analysis. It is important to note that fractionating each pI fraction into individual hydrophobic (HPLC) peaks is easy to do either in a multi-well format or by selective isolation of a range of peaks. Moreover, each of these sub-fractions contain the intact proteins that can be used for numerous other types of analysis such as Westerns, ELISA’s, LC/MS/MS, MALDI TOF, etc.
