Recently, the surface enhanced fluorescence technique has been helpful for several biological applications. Some researchers have reported the SEF of fluorophores such as dye molecules and QDs using dielectric spacers formed by several approaches. Herein, for the first time, we report the SEF spectra of CdSeS/ZnS alloyed quantum dots (QDs) that have high photostability, less toxicity, and high biocompatibility. Interestingly, the thin cytop layers formed by a simple approach are used as “dielectric spacers” between Au nanoparticles in the plasmon-active substrate (PAS) and the QDs. For comparison, the spectra of QDs dispersed directly on (i) bare PAS, (ii) bare glass substrate, and (iii) cytop coated glass substrate are also recorded. The enhancement factor is found to vary from one to two orders of magnitude. Since the SEF enhancement depends on the electric field intensity enhancement (EFIE) in the PASs, Finite Element Method (FEM) simulations are performed to estimate the EFIE and its dependence on the thickness of the cytop layer and size of the nanogap between nanoparticles in the PASs.