Electrospinning of type I collagen in 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) to fabricate a biomimetic nanofibrous extracellular matrix for tissue engineering was investigated. The average diameter of collagen nanofibers electrospun from 8% collagen solution in HFIP was 460 nm (range of 100-1200 nm). The as-spun collagen nanofibrous matrix was chemically cross-linked by glutaraldehyde vapor with a saturated aqueous solution and then treated with aqueous 0.1m glycine to block unreacted aldehyde groups. With vapor phase cross-linking for 12h, porosity of the collagen matrix decreased from 89% to 71%. The collagen nanofibrous matrix showed good tensile strength, even in aqueous solution. Effects on cytocompatibility, cell behavior, cell and collagen nanofiber interactions, and open wound healing in rats were examined. Relatively low cell adhesion was observed on uncoated collagen nanofibers, whereas collagen nanofibrous matrices treated with type I collagen or laminin were functionally active in responses in normal human keratinocytes. Collagen nanofibrous matrices were very effective as wound-healing accelerators in early-stage wound healing. Our results indicate that cross-linked collagen nanofibers coated with ECM proteins, particularly type I collagen, may be a good candidate for biomedical applications, such as wound dressing and scaffolds for tissue engineering.