Compared with free bacteria, immobilized bacteria have a higher oil degradation rate, making them highly promising for the bioremediation of offshore oil pollution. In our study, preparation conditions of modified Enteromorpha-immobilized bacterial agent were optimized by response surface method. A reactor was constructed and the effect of different diesel concentration and flow rate on the removal rate were investigate. The recyclability of modified Enteromorpha-immobilized bacterial agent was further studied. The result showed that, under the optimal preparation conditions, the removal rate of diesel in 5 days reached 90.39% and the mechanical strength reached a maximum value of 173.67g. When the inlet flow rate was not greater than 6.0mL/min and the concentration of diesel was less than 16.8g/L, the removal rate of diesel was guaranteed within 10.5h. After three cycles of diesel breakthrough, no obvious deterioration was noted in the diesel removal rate or fragmentation rate. This study provides a valuable insight into the practical application of immobilization technology to remove petroleum from seawater in the future.