Development of novel wound dressing with potent antibacterial activity is crucial for wound healing and tissue regeneration. In this work, we aim to prepare silver nanoparticles (AgNPs)-doped collagen-alginate (CA-AgNPs) biocomposite, which may possess antibacterial activity and be used as wound dressing. AgNPs were synthesized using NaBH4 as reducing agent and polyvinyl pyrrolidone as stabilizing agent. The formation of the AgNPs was confirmed by ultraviolet-visible spectrophotometer and transmission electron microscopy. Then, the as-prepared AgNPs were mixed with sodium alginate and collagen to obtain CA-AgNPs biocomposite. The CA-AgNPs biocomposite was fully characterized to verify the presence of AgNPs in the biocomposite. In vitro cytotoxicity assay illustrated that the CA-AgNPs biocomposite possessed negligible cytotoxicity at low AgNPs concentration. Furthermore, the antibacterial activity of the CA-AgNPs biocomposite was assessed against Staphylococcus aureus and Escherichia coli through agar diffusion method. Inhibition zone indicated that CA-AgNPs biocomposite possessed much higher antimicrobial activity than that of CA biocomposite, which strengthened with the increase in the AgNPs contents. Taken together, our finding suggested that the CA-AgNPs biocomposite showed strong potential as wound dressing.