Cellulose nanofibers (CNFs), chitosan, and silver nanoparticles (AgNPs) are widely used to enhance the active functions and antibacterial properties of wound dressings. This study was conducted to prepare CNF/AgNP-chitosan using a straight incorporation method and to assess its antimicrobial activity. CNFs were isolated from oil palm empty fruit bunches (OPEFBs) using the pulping method and acid hydrolysis. AgNPs were synthesized using a green synthesis method. The wound dressing was produced by mixing a 10% CNF solution in LiCl/DMAc and AgNP-chitosan in a glass plate with various ratios of CNF/AgNP-chitosan, i.e., 100:0, 80:20, 60:40, and 50:50. UV-visible and TEM analyses were conducted to confirm the formation of AgNPs and CNFs at the nanoscale. The results showed particles with an absorption wavelength of 435 nm and spherical shapes. Based on the calculation using ImageJ software, the diameters of CNFs were approximately 50 nm, and the lengths were several micrometers. FTIR was used to analyze the chemical bonding of AgNP-chitosan and the incorporated AgNP-chitosan in CNFs. Based on the XRD analysis, the presence of AgNPs did not affect the crystallinity of the CNFs. SEM images showed that the addition of AgNPs resulted in the stretching of CNF pores on the pads. Thermal degradation of the film increased with the addition of AgNP-chitosan by up to 40%. Antimicrobial tests and hemocompatibility tests showed that the formed CNF/AgNP-chitosan film successfully inhibited bacterial growth and was classified as a nonhemolytic material; thus, its potential as a wound dressing should be further studied.