Practical application to three-dimensional (3-D) tissue culture has been limited by the structural restriction of two-dimensional (2-D) nature of electrospun nanofiber mat. In this study, for constructing 3-D nanofibrous structure as real 3-D tissue engineering scaffold, we developed new fabrication process with silk fibroin (SF) by electrospinning and evaluated the features of this SF nanofiber scaffold (SFNS) through morphological and cell-culture analyses. Foam type of the SINS exhibited high porosity as well as large pores and its cell proliferation well occurred inside (inner spaces of pores), which makes this suitable for 3-D cell-culture scaffold.