New hybrid quantum dot (QD)/nanofibers have potential applications in a variety of fields. A novel fluorescent nanocomposite nanofiber material, consisting of CdS and zein has been fabricated through the electrospinning process. A detailed optimization was carried out to fabricate continuous and uniform nanofibers without beads or droplets. The synthesized hybrid nanofibers were characterized by various state-of-the-art techniques such as scanning electron microscopy, transmission electron microscopy (TEM), TEM-energy dispersive spectrometry, atomic force microscopy and confocal fluorescence micrography. The optimization process was carried out to fabricate fibers ranging from 200 to 450 nm in diameter. The electrical conductivity of the zein-CdS hybrid nanofiber substrates was tested. The potential use of the electrospun CdS-encapsulated nanofibrous scaffold as substrates for cell/tissue culture was evaluated with two different cell types, i.e. mesenchymal stem cells and fibroblasts. The results showed that the electrospun fibrous scaffolds could support the attachment and the proliferation of cells. In addition, the cells cultured on the fibrous scaffolds exhibited normal cell shapes and integrated well with surrounding fibers. The obtained results confirmed the potential for the use of the electrospun QD-encapsulated fluorescent nanofiber mats as scaffolds for tissue engineering.