Methods of preparing organosilane-functionalized regions on a substrate surface and more specifically fabricating patterned functionalized substrates suitable to be optically read, the methods generally comprising employing a vapor deposition process of an organosilane gas onto a lithographically patterned silicon surface followed by removal of the patterning media in a bath of organic solvents and ultrasonic excitation. The inventive methods provide optimized surface density of functional species while avoiding deleterious effects that can occur when lithographically patterned substrates are exposed to various gaseous species during the functionalization process.