In this work, the water-in-oil emulsion solvent diffusion method was developed as an alternative method for preparing cross-linked silk fibroin (SF) microparticles. Aqueous SF solution and ethyl acetate were used as water and oil phases, respectively. Influence of cross-linking on morphology and thermal stability of the SF microparticles were investigated. Genipin was used as a cross-linker. The SF microparticles had spherical shape with smooth surface when Span80 was used as the oil-soluble emulsifier. The SF in aqueous solution was cross-linked with genipin before microparticle formation. Internal morphology of the SF microparticles observed on broken microparticles showed as sponge. The cross-linking induced SF conformational changes from random coil to β-sheet form. Porosity of the SF microparticle matrices decreased and thermal stability increased when the SF was cross-linked and the degree of cross-linking was increased. © 2011 Curtin University of Technology and John Wiley & Sons, Ltd.