Microbial column flotation using Thiobacillus ferrooxidans was applied for desulfurization of Pittsburgh coal of CWM (Coal-Water Mixture) size between 38 mu m and 75 mu m. The coal contained ferrous ion which would interfere separation of pyrite from coal by microbial flotation. The wash-out of ferrous ion with 0.5N HCl solution enabled pyrite removal from coal. The coal was divided into two parts, the small-size coal between 38 mu m and 53 mu m, and the large-size coal between 53 mu m and 75 mu m. The pyritic sulfur content was decreased from 2.88% of the feed coal to 0.98% of the product coal for the large-size coal and from 2.77% of the feed coal to 1.12% of the product coal for the small-size coal by microbial flotation. The decrease was based on removal of liberated pyrite particles (between 20 mu m and 70 mu m). However, the fine particles (less than 20 mu m) could not be removed even though the pyrite particles were liberated from coal particles. The microbial column flotation was more effective for desulfurization of the large liberated pyrite particle than that of the small. It was not effective for desulfurization of the locked pyrite particles that were buried in coal particles. Both the pyrite liberation from coal and its particle size are important factors for the pyrite removal by microbial column flotation.