In this presentation, we propose a fabrication method of PDMS stencil to apply into a localized culture of NIH/3T3 cells. PDMS stencil was fabricated by nitrogen gas blowing and soft lithography from preparing SU-8 master mold by photolithography. PDMS stencil pattern was production of the circle size 20 to 500 μm. In the culture test of PDMS stencil, a stencil was placed on a glass disk. The NIH/3T3 cells were successfully cultured into micropatterns by using the PDMS stencil. The results showed that cells could be cultured into micropatterns with precisely controlled manner at any shapes and specific size for bioscience study and bioengineering applications.