Enzyme-linked immunosorbent assays (ELISA) is one of the most commonly used immunoassays for detection and quantification of chemical and biological molecules (antigens; mainly proteins and polypeptides) in the biotechnology industry, and is becoming more and more important in clinical diagnostics, food safety testing, and environmental monitoring. Recently, research attention has been focused on developing a miniaturized and highly multiplexed microarray format. In this study, instead of using the 96-well microtiter plate, we proposed to utilize the micromolding in capillaries (MIMIC) technique for fabrication of the microarray-based ELISA. By modifying the conventional ELISA procedures, we were able to pattern the antiboby and antigen on the substrate to perform ELISA process. This technique provides large-area patterning to fabricate microarray-based ELISA at a fast, low-cost and reliable fashion. Moreover, the design of the feature size and selection of the substrate is relatively flexible.