In this study, we characterized strain F9 and evaluated the interaction between strain F9 and hematite by scanning electron microscopy (SEM), Fourier transform infrared spectrophotometry (FTIR), zeta potential, flotation, and other methods. The results showed that strain F9 belongs to Serratia marcescens. This brevibacterium had CH₂, CH₃, and hydroxyl groups on its cell wall, which imparted a strong hydrophobic and negative charge. Adsorption of strain F9 reduced the zeta potential of the hematite surface and increased the hydrophobicity of the hematite surface, thereby generating hydrophobic hematite agglomerates. At least four groups on strain F9 interacted with the hematite surface, which contributed to chemical interactions of carboxylic groups and hydrophobic association among hydrophobic hematite particles. The possible use of strain F9 as a bio-collector for hematite flotation was proved.