Surface biofunctionalization of group-III nitride semiconductors has recently attracted much interest due to their biocompatibility, nontoxicity, and long-term chemical stability under demanding physiochemical conditions for chemical and biological sensing. Among III-nitrides, aluminum nitride (AlN) and aluminum gallium nitride (AlGaN) are particularly important because they are often used as the sensing surfaces for sensors based on field-effect transistor or surface acoustic wave sensor structures. Patterned self-assembled monolayer (SAM) templates are composed of two types of organosilane molecules terminated with different functional groups (amino and methyl), which were fabricated on AlN/sapphire substrates by combining photolithography, lift-off process, and self-assembly technique. Clear imaging contrast of SAM micropatterns can be observed by field emission scanning electron microscopy (FE-SEM) operating at a low accelerating voltage in the range of 0.5–1.5 kV. In this work, the formation of green fluorescent protein (GFP) antibody microarrays was demonstrated by the specific protein binding of enhanced GFP (EGFP) labeling. The observed strong fluorescent signal from antibody functionalized regions on the SAM-patterned AlN surface indicates the retained biological activity of specific molecular recognition resulting from the antibody–EGFP interaction. The results reported here show that micropatterning of organosilane SAMs by the combination of photolithographic process and lift-off technique is a practical approach for the fabrication of reaction regions on AlN-based bioanalytical microdevices.