Determination of 135Cs concentration and 135Cs/137Cs atomic ratio is of great importance in characterization of radioactive waste from decommissioning of nuclear facilities. In this work, an effective analytical method was developed for simultaneously determination of 135Cs and 137Cs in different types of waste samples (steel, zirconium alloy, reactor coolant, ion exchange filter paper and spent ion exchange resin) by coupling AMP-PAN, AG MP-1M and AG 50 W-X8 chromatographic separation with ICP-MS/MS measurement. Decontamination factors of 7.0 × 106 for Co, 6.0 × 106 for Ba, 4.2 × 105 for Mo, 3.2 × 105 for Sn and 2.1 × 105 for Sb were achieved using the chemical separation procedure. The overall chemical yields of cesium were higher than 85%. A detection limit of 3.1 × 10−14 g/g for 135Cs was achieved for 0.2 g stainless steel sample or spent resin. The developed method was validated by analysis of standard reference materials (IAEA-375) and successfully applied for analysis of zirconium alloy, steel, ion exchange filter paper and spent ion exchange resin from nuclear power reactors. The obtained 135Cs can be used to evaluate the long-term environmental impact and provide useful information for waste disposal. The measured 135Cs/137Cs ratio in reactor coolant, as a characteristic information, might be useful for source identification and localization of leaked fuel element. The neutron flux of the leaked fuel element can be estimated based on the measured 135Cs/137Cs atomic ratios in the reactor coolant water. The developed method is simple and rapid (8 samples/day) for the determination of 135Cs concentrations and 135Cs/137Cs ratios in various waste samples from nuclear decommissioning.