Because of their wide applications in manufacturing life science and medicine, nanoparticles (NPs) are becoming potential limiting factors for agricultural plant growth and productivity. Thus, development of some low cost and low risk methods for detoxifying or alleviating cytotoxicity of NPs on crops would be a proactive attractive solution. In this study, we investigated the ameliorating effect of He-Ne laser illumination of seedlings on cytotoxicity of cadmium sulfide nanoparticles (CdSNPs) in tall fescue seedlings. Physiological parameters and biochemical characteristics of 20-d-old seedlings with different times of laser illumination and graded doses of CdSNPs treatment, either alone or in combination, were measured. Cadmiun sulfide nanoparticles exhibited pronounced inhibitory effect on shoot and root growth, respectively. Accumulation of CdSNPs in plant root cells resulted in oxidative stress as detected by the formation of reactive oxygen species (ROS), which aggravate the cytotoxicity of CdSNPs on seedlings. Treatment of seedlings with CdSNPs alone also reduced antioxidant concentration, phytochelatins (PCs) content, enzymatic activities of phytochelatin synthase (PCS), and antioxidant enzymes, such as CAT, APX, GR, and POD, except SOD. However, He-Ne laser illumination prior to CdSNPs exposure alleviated the inhibitory effect on seedling growth and development through activation of tall fescue antioxidant systems due to induction of antioxidants biosynthesis and antioxidant enzyme activities. Furthermore, PCs and GSH induced by laser was also demonstrated to be involved in decreasing the cytotoxicity of CdSNPs through binding free cadmium ions release from CdSNPs. The interactive mechanisms between He-Ne lasers, plant antioxidant systems, and PCs biosynthesis pathways need be further investigated.