Enzyme-linked immunosorbent assay (ELISA) is widely used in medical diagnostics and fundamental biological research due to its high specificity and reproducibility. However, the traditional 96-well-plate based ELISA still suffers from several notable drawbacks, such as long assay time (4-6 hours), burdensome procedures and large sample/reagent volumes (∼100 μl), which significantly limit traditional ELISA's applications in rapid clinical diagnosis and quasi-real-time prognosis of some fast-developing diseases. Here, we developed a user friendly glass capillary array based microfluidic ELISA device. Benefiting from the high surface-to-volume ratio of the capillary and the rapid chemiluminescent photo-imaging method with a commercial camera, our capillary based ELISA device significantly reduced the sample volume to 20 μL and shortened the total assay time to as short as 16 minutes (including detection time), which represent approximately 10-fold and 5-fold reduction in assay time and sample volume, respectively, in comparison with the traditional plate-based method. Furthermore, through the double exposure method, a nearly 10-fold increase in the detection dynamic range was achieved over the traditional well-based ELISA. Our device can be broadly used in rapid biochemical analysis for biomedicine and research/development laboratories.