Bacterial cellulose (BC) tissue engineering scaffold, possessing nano-fiber, an ultrafine 3-dimensional network and proper porosity, has been prepared by static culture of G. xylinus. It has been characterized by scanning electron microscope (SEM), atomic force microscope (AFM), and light microscope. The pore size of BC is 0.6 to 2.6 micrometers (mum) and the width of cellulose ribbon is 50 to 80 nanometers (nm). By measuring their dry weight and wet weight, the porosities of air-dried BC and freeze-dried BC have been determined to be around 70% and 90% respectively; the excellent permeability of freeze-dried one has also been found to be better than PVC and PE film. The results show that it has extremely fine network of nano-structure, and the water vapor permeability is quite excellent. Cultured with fibroblasts (FBs) and chondrocytes respectively and subcutaneous transplantation of FBs-BC composition, we find that BC is suitable for cell attachment and proliferation, and shows better cell compatibility. It is expected to become a potential tissue engineering scaffold.