A method of fabricating a composite PDMS microstructure includes a defining step, a depositing step and an etching step. The defining step is performed for defining a patterned area having a mono-molecule with a thiol group on a PDMS substrate, and the mono-molecule with the thiol group is in liquid phase. The depositing step is performed for placing the PDMS substrate having the mono-molecule with the thiol group into a vacuum chamber within an activation time so as to deposit one Au atom on the patterned area of the PDMS substrate by a vacuum coating process. The etching step is performed for cleaning the PDMS substrate using water, and thus the Au atom can be selectively retained on the patterned area of the PDMS substrate.