Sol-gel derived bioactive glass nanoparticles (BGNs) are fascinating materials for bone regeneration. In the literature, it can be found that their specific surface area and their calcium content (Ca/Si ratio) are the two key parameters impacting strongly the particles bioactivity. Nevertheless, in most studies, in vitro bioactivity tests are performed on series of materials where both the composition and the specific surface area are varied. It is thus difficult to unravel the effect of each parameter independently. In this study, spherical and monodispersed BGNs with different Ca/Si ratio and similar specific surface area have been synthesized by a modified Stöber method in order to specify the impact of the calcium content only. The mineralization studies performed in simulated body fluid showed that the bioactivity increases with the amount of calcium incorporated in the glass matrix. However, this effect is not significant in the composition interval studied (7-15 %mol of CaO). Such a result proves that the effective Ca/Si ratio is not the major parameter which affects the bioactivity of sol-gel binary BGs. In vitro biocompatibility assessment during 3 and 7 days using human Mesenchymal Stem Cells in contact with the sample showing the fastest mineralization proved its non-cytotoxicity. Hence, biomedical applications can be intended for this sample.