Abstract: Polyhexymethylene Phenformin hydrochloride (PHMB) and polystyrene sulfonate (PSS) were alternately deposited on the surface of cotton fabric via electrostatic self-assembly which endowed the fabric with antibacterial ability. The surface morphology of assembled fibers was revealed via scanning electron microscopy. The surface of assembled fibers became rough, proving that PHMB/PSS were coated on the fiber. Zeta potential of the fibers and dyeing color depth showed that the distinct "layer-by-layer alternating oscillation" along with the change of deposition materials on fiber surface, which helped to investigate the assembly mechanism. The bacterial inhibition rate of self-assembled cotton fabrics reached 99.51%, showing the excellent anti-bacterial property. In addition, the assembled cotton also had a good washing fastness.