Patterned microarrays of antibodies were fabricated and tested for their ability to bind targeted bacteria. These arrays were used in a series of bacterial immunoassays to detect E. coli 0157:147 and Renibacterium salmoninarum (RS). Microarrays were fabricated using microcontact printing (muCP) and characterized using scanning probe microscopy (SPM). The high-resolution SPM imaging showed that targeted bacteria had a higher binding selectivity to complementary antibody patterns than to unfunctionalized regions of the substrate. Additional studies indicated a significant reduction in binding of bacteria when the microarrays were exposed to nonspecific bacteria. These studies demonstrate how protein microarrays could be developed into useful platforms for sensing microorganisms.