This work is concerned with synthesis and characterization of electrospun nanofibers for advanced drug delivery and cell culturing. Poly--e-caprolactone (PCL), composed of different inclusions such as gentamicin, plasmid DNA, and gelatin, is used to fabricate nanofibers using electrospinned for drug delivery and cell culturing. The plasmid DNA enhanced green fluorescent protein (EGFP) with cyromegalovirus (CMV) promoter (PCMVb-GFP) was ampkufued with Escherichia coli (E. coli). The plasmid DNA enhanced green fluorescent protein (EGFP) with cytomegalovi.i1.1s (CMV) promoter (PCMVb-GFP) was amp lified with Esche;ichia coli (E. coli). PCL was chosen because it has been approved by the Food and Drug Administration (FDA) in implantable material applications used iii the human body, and it has biocompatible and biodegradable material. which plays a critical role in tissue engi.i1eering. wound heali.i1g, and drug delivery. The main reason for using electrospun nanofibers in this study is because it has high surface area. porosity, and penneability. Sca1ming electron microscopy (SEM) images show that the nanoscale fiber structures have a diameter ranging of 50 to 250 mn with some bead fonnations. The cytotoxicity study revealed that PCL nanofibers are not toxic, and cell viability was above 70%. An energy dispersive x-ray spectroscopy (EDS) was used iii order to confirm the ratio of the gelati.!1 component iii the fibers. To visually express the inclusio n of gentamicin iii the fibers a fourier transform infrared (FTIR) spectroscopy was used. Differential scanning calorimetry (DSC) and thermal gravimetric analysis (TGA) techniques were also applied on the prepared sample. Overall, this study showed that PCL is a good candidate for several biomedical applications and may open up new possibilities for partiicularly DNA, gene, nerve cell myelination and drug delivery purposes.