The gelatin film has great potential in biomedical applications, especially in wound healing. The combination of gelatin films and stem cells could further accelerate the skin regeneration. Although polysaccharide modification can improve the mechanical property and biological activity of gelatin films, information about the stability and cellular affinity is still limited. This study investigated the influence of polysaccharides on the stability and cellular affinity of gelatin films. Two kinds of gelatin-polysaccharide composite films, including gelatin-hyaluronic acid (G-HA) and gelatin-chitosan (G-CS), were prepared in this study. It was found that G-HA composite film had better short-term and long-term stability compared with G-CS composite film. And G-HA composite film also had better biological safety than G-CS film. Moreover, the surface of G-HA composite film supported the adhesion and growth of human umbilical cord Wharton's jelly-derived mesenchymal stem cells (WJ MSCs) better than G-CS film surface. These data illustrated that G-HA composite film has better stability and cellular affinity compared with G-CS film, which could be considered a promising delivery system of stem cells for further in vivo studies. Therefore, this work would be very helpful to optimize the preparation of gelatin-polysaccharide composite films.