The field of wound healing has seen an increase in research activity in wound care and hydrogel-based dressings have been targeted as a solution for these applications. Hydrogels with silver nanoparticles can present many advantages for this field. However, if the aggregation and sterilization of this product have not been carefully considered, the effectiveness or use could be limited. Therefore, in the current study, a hydrogel-based wound dressing membrane was developed using polyvinylpyrrolidone (PVP), polyethylene glycol (PEG), agar, and carboxymethyl cellulose (CMC). Silver ions (Ag+) were dispersed in the polymer matrix and its reduction with formation of a hydrogel and silver nanoparticles was performed using 60Co gamma irradiation to enhance the dressings antimicrobial properties. The resulting hydrogel presented a high degree of swelling and a good size control of silver nanoparticles. The incorporation of AgNPs was confirmed via Raman spectroscopy and the samples presented no signs of toxicity in vitro as assessed using an elution assay with neutral red uptake as the cytotoxic end point. Membranes were tested in vivo using a full thickness defeat model in rabbits. Postmortem histopathological analysis indicated that the use of the hydrogel membranes that incorporated AgNPs had a stimulatory action on wound healing as evidenced by a high intensity of fibroblasts and neovascularization in the tissue, which promoted a faster healing process when compared to the untreated wounds. We demonstrate the possibility of producing a hydrogel with good size control of AgNPs, which can also be directly sterilized within the formation of this material via gamma irradiation. Furthermore, the mechanism of hydrogel healing, in vivo, with silver nanoparticles was found to have a direct correlation of silver nanoparticles with in vitro cell results.