The application discloses a kit for specifically detecting creatine kinase isoenzymes, which belongs to the technical field of creatine kinase detection. The kit includes reagent R0, reagent RM and reagent R1; reagent R0 includes buffer, sodium chloride, protective agent, preservative, creatine kinase isoenzyme B subunit antibody; reagent RM includes buffer, coated muscle Acid kinase MM isozyme-coated magnetic microspheres, protective agent, surfactant, stabilizer, preservative; reagent R1 includes buffer, alkaline phosphatase-coated creatine kinase MM isozyme-labeled antibody, Protective agent, surfactant, stabilizer, preservative. The kit of this application has high sensitivity, good specificity, high linear range and HOOK resistance, and can detect CKMM in dry filtered blood film, serum or urine samples, and has a wider scope of application.