The objective of this research was to investigate the catalytic role of nitrogen-doped carbon nanotubes (NCNTs) in a bioreactor that contained Acidithiobacillus ferrooxidans (A. ferrooxidans) and waste printed circuit boards (WPCB). Factors that could reflect the results, such as pH, oxidation-reduction potential (ORP), Fe²⁺, and leaching rate of copper were detected and calculated in the bioleaching process. The results demonstrated that a certain amount of NCNTs had good compatibility with A. ferrooxidans, and it had the best consistency when added to 0.04g NCNTs in 50 ml solution. NCNTs could promote nearly 10% of copper leaching rate from WPCB by A. ferrooxidans compared with the blank group. When the NCNTs were treated by acid solution, its ability to promote the leaching rate of copper had a certain degree of decline (but not obvious), and it was about 2% lower than the group in which the NCNTs were not treated with acid. These phenomena could expand the application of NCNTs, and these could provide a basis for the next industrial application of A. ferrooxidans.