Recently, menstrual blood has been identified as an easily accessible, refreshing and renewable stem cell source. The aim of this study was to evaluate proliferative potential of MenSCs in silk fibroin/gelatin scaffold compared with 2D culture . Menstrual blood samples (2-5 ml) were collected from 5 healthy female volunteers with the age range of 20-30 years. Then, MenSCs were isolated of obtained samples by discontinuous density gradient centrifugation. After assessment of imunophenotypic properties of cultured cells, MenSCs were seeded in the fabricated silk fibroin /gelatin scaffolds that were characterized using scanning electron microscopy (SEM), FT-IR and universal testing machine. Then proliferative capacity of MenSCs in silk fibroin/gelatin scaffold was compared with 2D condition using propidium iodide DNA quantification assay. Flowcytometric analysis illustrated that MenSCs typically express the surface antigens associated to mesenchymal stem cells whereas the expression of Hematopoietic stem cells markers in these cells was negative. Based on SEM images, MenSCs were attached, penetrated and distributed well in the scaffold. While proliferation rate of MenSCs was significantly higher level in scaffolds than that in two dimensional (2D) culture (P < 0. 01). Based on accumulative data, MenSCs are a unique cell population with higher proliferation capacity . It seems that silk fibroin /gelatin scaffold could be viewed as a novel, safe, natural and accessible construct for tissue engineering.