An In-Gel Assay for Protein Tyrosine Phosphatase Activity: Detection of Widespread Distribution in Cells and Tissues

states. During the past few years, however, it has bepolymer) is incorporated into gels prior to polymeriza- come apparent that PTPases may participate more action. Following electrophoresis, the sodium dodecyl tively in signaling pathways. In many cases the activity sulfate is removed; the proteins are fully denatured by of tyrosine kinases is itself regulated by tyrosine phossoaking gels in 6 M guanidine hydrochloride and then phorylation, and so the removal of phosphates from renatured by incubation in buffers containing 0.04% tyrosine kinases by PTPases may lead to either their Tween 40 and high concentrations of reducing agents. activation or inhibition. During the past few years a Protein tyrosine phosphatase activity is detected in large number of PTPases have been isolated, cloned, autoradiographs of dried gels as regions from which and sequenced (3‐5). Two distinct groups of PTPases the 32 P has been selectively removed. Electrophoresis have been identified. One group is transmembrane proof known cytoplasmic protein tyrosine phosphatases teins, with extracellular domains that resemble recepindicates activity at the predicted molecular weights. tors, although ligands for these have generally not been As little as 10 pg of some cytoplasmic phosphatases is identified. The other group of PTPases is cytoplasmic. detectable. However, transmembrane tyrosine phos- Many of these reveal domains, such as src homology 2 phatases, such as CD45, are detected only at very high (SH2) domains, suggestive of complex interactions or protein loadings in this assay. Electrophoresis of modes of regulation (3‐5). In addition, within this latwhole cell lysates indicates multiple bands of tyrosine ter category there is a subgroup of phosphatases that