Abstract: ~(135)I-creatine phosphokinase-MM(~(125)I-CPK-MM) was prepared by ~(125)I-labelled Bolton-Hunter reagent(HPNS). Iodinating conditions of HPNS and its conjugation to protein were studied. ~(125)I-CPK-MM with immune activity was obtained and used to establish the ~(125)I-CPK-MM radioimmunoassay method by the General Hospital of PLA. ~(125)I-CPK-MM in PBS-G solution containing 0.015mol/1 ethyl mercaptan at 4-10℃ can be used for one month.