The suitability of hemp fibres as green support material for immobilization of Acidithiobacillus ferrooxidans cells in ferrous iron bio-oxidation process was investigated. A series of experiments were carried out in shake flasks and bioreactor for both cell immobilization and iron bio-oxidation. In flask experiments, five successive batch colonization cycles resulted in 6-fold increase in ferrous iron biooxidation rate. Evaluation of immobilized cells activity through two approaches as wet and dry storage demonstrated the higher activity of wet stored bacteria than dry ones after a period of two months. The dry stored bacteria lost almost all their activities after eight weeks. Whereas, the activity of wet stored samples was 30% at the same time. In bioreactor experiments, use of a packed-bed bioreactor with hemp fibres, the maximum ferrous iron bio-oxidation rate of 2.46 kg m -3 h -1 was obtained at dilution rate of 3.97 h -1 , and retention time of 0.25 h. Our experimental results could pave a new platform for the application of hemp fibre as an inexpensive and environmentally friendly support material in bacterial cell immobilization, especially in ferrous iron bio-oxidation process using acidophilic bacteria.