The activation energy associated with the electrophoretic migration of an analyte under given electrolyte conditions can be accessed through the determination of the analyte electrophoretic mobility at various temperatures. In the case of the electrophoretic separation of polyelectrolytes in the presence of an entangled polymer network, activation energy can be regarded as the energy needed by the analyte to overcome the obstacles created by the separating network. Any deformation undergone by the analyte or the network is expected to induce a decrease in the activation energy, in this work, the electrophoretic mobilities of poly(styrenesulfonates) (PSSs) of various molecular weights (M-r 16x10(3) to 990x10(3)) were determined in entangled polyethylene oxide (PEO) solutions as a function of temperature (in the 17-60 degreesC range) and the PSS activation energies were calculated. The influences of the PSS molecular weight, blob sizes ib Of the separating network (related to the PEO concentration), ionic strength of the electrolyte and electric field strength (75-600 V/cm) were investigated. The results were interpreted in terms of analyte and network deformations and were confronted with those previously obtained for DNA migration in polymer solutions and chemical gels. For a radius of gyration R-g<(b) the activation energy increases with the PSS molecular mass, while the reverse is true for R-g>zeta (b), suggesting PSS and network deformations in the latter case. Increasing ionic strength resulted in an increase in the PSS activation energy, because of the decrease of their radii of gyration, which makes them less deformable. Finally, the activation energies of all the PSSs are a decreasing function of field strength and at high field strength tend to reach a constant value close to that for a small molecule.