As an excellent biocompatible and biodegradable protein polymer, silk fibroin (SF) has found wide applications, particularly serving as therapeutic agent for tissue-engineering applications, on which both post-spin treatment and sterilization processing are crucial to drug-loaded matrices. To find a safe, effective and appropriate post-spin treatment and sterilization approach for drug-loaded biomaterial matrices is one of the major problems in the field of tissue engineering at present. In this work, a simple, safe and effective approach skillfully integrating post-spin treatment with sterilization processing was developed to drug-loaded SF nanofibrous matrices. Electrospun SF nanofibrous matrices from its aqueous solution were post-treated with 75% ethanol vapor. C-13-NMR and WAXD analysis demonstrated that such post-spin treatment rendered the structure of SF nanofibrous matrices transform from the silk I form to the silk II form. Furthermore, biological assays suggested that as-treated SF nanofibrous matrices significantly promoted the development of murine connective tissue fibroblasts. Skillfully integrated with novel sterilization processing, 75% ethanol vapor treatment could be a potential approach to designing and fabricating diverse drug-loaded SF nanofibrous matrices serving as therapeutic agents for tissue-engineering applications in that it can effectively protect the drug from losing compared with traditional post-spin treatment and sterilization processing. (C) Koninklijke Brill NV, Leiden, 2012