In this chapter we present the results of the photoluminescent and optical investigations of the influence of cation vacancy-related defects on CdSe/ZnSe quantum dot organization. Selfassembling growth was achieved under molecular beam epitaxy with subsequent annealing step. Number of cation vacancies was controlled by the intensity of the emission band connected with complex that includes cation vacancy and shallow donor. For the first time it is shown that increase of number of cation vacancy related defects results in the reduction of potential fluctuations in the QD layer. In this case a relatively uniform dense array of QDs with shallow localization potential is organized. It is proposed that generation of cation vacancies during the growth suppresses both Cd segregation and Cd surface diffusion as well as facilitates Cd/Zn interdiffusion. Interdiffusion process is proved by the changes in the photoluminescence and optical reflection spectra of ZnSe layers. It is showned that Cd/Zn interdiffusion can play an important role in CdSe/ZnSe intermixing during the QD formation at least under such growth conditions which can stimulate generation of cation vacancies.