The invention discloses a testosterone detection reagent based on a microparticle chemiluminescence immunoassay technology; the testosterone detection reagent comprises the following components: 0.002%-0.01% of paramagnetic microspheres, 0.4 mug / ml-1 mug / ml of acridinium ester labeled testosterone antibodies, a composite testosterone release agent, a luminescence liquid A, a luminescence liquid B, 0ng / dl-1500ng / dl of a testosterone calibrator and a cleaning fluid with a certain concentration; and the testosterone release agent is a mixture of dihydrotestosterone, danazol and heparin sodium. The testosterone detection reagent has the advantages of simple operation, high sensitivity, fast detection efficiency, low cost, easy automation and the like, and by combination of the selected composite release agent and different binding globulin, combined-state testosterone in serum can be fully released, so that the content detection of total testosterone is more accurate.