The present study aimed to investigate the effect of silver nanoparticles (Ag-NPs) as an anti-ethylene agent on in vitro microtuberization in potato cv. Spunta. The concentrations; 0, 0.5, 1, 2, 4, 8, and 16 mg/l of Ag-NPs or silver thiosulfate (STS) were used to determine their influence on microtuberization. The results of Ag-NPs treatments showed that, the average number of microtubers/jar was elevated and reached the highest level (14.9 microtubers/jar) which represents about 83.95% over the control treatment (8.1 microtubers/jar) using the medium containing (1 mg/l) Ag-NPs. While the highest level (10.9 microtubers/jar) was recorded using (8 mg/l) STS, which is about 34.56% over the control treatment. Thus, these results confirmed that the low level of Ag-NPs (1 mg/l) was the most effective for inhibiting ethylene action on microtuberization in potato cv. Spunta. In addition, these findings complement and agree with those obtained in our previous study using the cv. Desirѐe (Ibrahim et al. 2018a). Therefore, we report here that an efficient genotype-independent protocol for microtuberization in potato has been established. In addition, this developed protocol using Ag-NPs is important for enhancing potato microtuberization technology at the mass-production level in Egypt. Furthermore, these results are of great importance in plant physiology and nanobiotechnology research.