When you see what appears to be proteins of identical MW in a 2D gel but appear to have many isoforms that differ in their isoelectric point (pI), you might want to assume these possibilities: 1) a protein with multiple glycosylation sites that feature sialylated oligosaccharides (these are negatively charged sugars) 2) a protein with multiple phosphorylation sites which are phosphorylated In the case of (1), you can possibly pre-treat the protein (or protein mixture) with a deglycosylating enzyme (perhaps PNGase F, also called N-glycanase) or with a de-sialylating enzyme (neuraminidase) and see if the multiple spots disappear into one spot on the 2-D gel. In the case of (2), you should be able to dephosphorylate the protein in some way. There are protein phosphatase specific for serines/threonine type phosphorylation (PP1, PP2a, PP2b, etc.) and some specific for tyrosine phosphorylation, but these are often quite expensive, and perhaps insufficient as a preparative treatment. A less expensive treatment is with alkaline phosphatase, but that may not be specific for protein. It is reported to dephosphorylate peptides, but as for intact proteins of fairly average-to-large MW, it might require extensive denaturation of the polypeptide, including having urea present while the alkaline phosphatase (ALP) hydrolyzes the phosphates. There are urea-resistant and heat-stable ALP enzymes reported in the literature, in particular in increased activity from the neutrophils of pregnant women. Other kinds of dephosphorylation include chemical means (beta elimination reactions to form alkenes). SMH --------------------------------- Everyone is raving about the all-new Yahoo! Mail beta.