In the field of tissue engineering, materials such as polycaprolactone (PCL), collagen and gelatin (Gel) are widely used in the production of scaffolds to enhance cell growth and support. The aim of this study was to produce and characterize electrospun scaffolds of PCL/Gel coated with tilapia skin collagen biofilm and to investigate their application in tissue regeneration. The methodology included the production of scaffolds using the electrospinning technique, while collagen was extracted from tilapia skin. The electrospun fibers were characterized through contact angle, scanning electron microscopy and fiber diameter measurement with ImageJ. The viability with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT assay) and adhesion with fluorescein diacetate, propidium iodide and 4’,6-diamidino-2-phenylindole of the mesenchymal stem cells were evaluated after two days of cultivation in the biomaterials. The results showed that the electrospun scaffolds had long, continuous fibers without beads, measuring approximately 500 nm in diameter. Contact angle testing indicated hydrophilicity with a value of 47.3 ± 12.7°. Fluorescence microscopy with cell staining assay demonstrated the attachment of the cells to the materials. In the MTT assay, a statistically significant difference was noted between the PCL/gel group and the control group, suggesting a higher number of viable cells in this scaffold. The other materials increased, but not statistically significantly, the stem cell viability, with absorbance values of 0.201 ± 0.015 for the cells cultivated on the tissue culture plate (control), 0.257 ± 0.050 for the cells on collagen, and 0.240 ± 0.015 in the PCL/gel/collagen group. In conclusion, all the materials tested maintained cell viability and were not cytotoxic. Therefore, the electrospun scaffolds of PCL/gelatin with or without tilapia collagen are suitable for cell therapy and tissue regeneration applications.