Silver nanoparticles (AgNPs) loaded on the cotton ber can endow the ber with good antibacterial activity, but the AgNPs on the surface of cotton ber will leach out with the increase of washing times, which will not only greatly reduce the antibacterial properties of cotton ber, but also cause pollution to the environment. In this paper, carboxyl groups were introduced into cotton ber by three methods, namely selective oxidation, butane tetracarboxylic acid (BTCA) grafting and polyacrylic acid (PAA) adsorption. The silver ions (Ag + ) were adsorbed on the cotton bers through electrostatic attraction, and after reduction, the AgNPs were loaded onto the cotton bers, and the AgNPs distribution on the ber and its effect to the washing resistance of AgNPs were studied. The results showed that the selective oxidation and the BTCA grafting can introduce carboxylic groups into the interior of cotton bers while the PAA adsorption only imports carboxyl groups on the surface of the ber. As a consequence, AgNPs were formed inside of the cotton ber when the selective oxidation and the BTCA grafting were employed, while AgNPs can only be loaded on the surface of the ber by the PAA adsorption method. AgNPs inside of the ber have high energetic adherence, resulting in silver-cotton fabric nanocomposites with strong slow-release property and long-term antibacterial ability. The fabric nanocomposites prepared by selective oxidation and BTCA grafting exhibited high AgNPs retention rate and excellent antibacterial activity even after 80 washing cycles.