Much research over the past five decades has focussed on the repair and replacement of bone. Recently, the research focus has shifted to nanotechnology since it provides a platform from which to alter and possibly improve materials’ properties. In this study we have made use of previously developed electrospun biphasic nanoscaffolds to culture osteoblast cells on, and investigate specific responses of the cells towards the scaffolds. Osteoclast-like cells and osteoblast cells were cultured separately on the nanoscaffolds and the proliferation, adhesion and cellular response were determined. In this study, the mineralisation of the osteoblast cells was observed in a time study. The intracellular calcium ion concentration and nitric oxide concentration were determined in vitro while the cells were proliferating on the scaffolds. The expression of endothelial and inducible nitric oxide synthase was determined immunohistochemically. Quantitative data were obtained from fluorometer studies. Qualitative data was supplied by light- and fluorescent confocal microscopy. During studies with microscopy, a minimum of five representative images from each sample were captured. The cells showed increased mineralisation over time. An increase in intracellular Ca2+ was not observed when compared to the controls. However, an increase in intracellular nitric oxide formation was detected. Expression of endothelial nitric oxide synthase but not inducible nitric oxide synthase was detected in vitro. From the results we can conclude that the scaffolds are biocompatible and conducive to healthy cell growth and differentiation and could possibly be applied in non-load-bearing bone regeneration and repair applications.