Bacterial samples were prepared taking cloth strip as carriers infected by bacillus subtilis var. niger spores. Bacterial carriers were disinfected by spraying chlorine-containing disinfectant coupled with heat effect. Orthogonal experiments were carried out taking available chlorine concentration, spraying quantity of disinfectant, temperature and sterilizing time as influencing factors. The results showed that spraying quantity 0.12 g/cm2of disinfectant containing 5000 mg/L available chlorine, bacillus subtilis var. niger spores on the carrier with 1×106cfu per sample could be killed completely (100%) in 60°C thermostat for 15 min. At the same time, contrastive experiments indicated that there were still 1100 cfu of residual bacteria on the carrier which had been sprayed disinfectant containing 5000 mg/L available chlorine and put into 25°C (room temperature) thermostat for 30 min. While no killing efficiency was observed when putting bacterial carrier into 60°C thermostat for 30 min, if taking standard hard water instead of disinfectant.