Novel nanostructured microcantilever (MC) surfaces were developed by modifying the active side of the MCs with aluminum oxide nanoparticles (AONP) for purposes of enhancing sensitivity in nanomechanical-based sensing. Uniform layers of AONP were spin coated and chemically immobilized on the surfaces of MCs with tetramethoxysilane (TMOS) as a cross-linker. Optimization studies on MC modification were performed for better surface uniformity and higher surface area, based on scanning electron microscope (SEM) images. The AONP-modified MC array (MCA) were subsequently functionalized by being immersed in parallel configured capillaries filled with different reagents for immobilizing chemical or biological receptors onto the MC surfaces. A MCA prepared for chemical sensing was exposed to the samples made of headspace vapor of different volatile organic compounds (VOCs). The characteristic response signatures for each gas phase VOC analyte showed substantial diversity. Immersion time in the capillary and the chemical nature of the reagents used for functionalization were both optimized to achieve the highest sensitivity and long-term reproducibility in nanomechanical responses to the test analytes. A second MCA functionalized with two different immunological receptors was prepared and exposed to three biological analytes in the liquid phase, with a highly selective response obtained for each analyte. Fluorescence microscope images and FT-IR spectra were used in this work to validate the controlled, variable chemical nature of the MC surfaces.