We have developed an enzyme-linked immunosorbent assay (ELISA) chip based on a digital microfluidic platform, in which droplet generation and manipulation are implemented using the electrowetting on dielectrics (EWOD) effect. First, droplet transportability was experimentally tested on silicon test chips with different control electrodes for optimizing the EWOD-based droplet manipulation, and then tested on transparent indium-tin-oxide (ITO) chips. The latter were then used for the development of an ELISA chip, on which we carried out immunoassay with rat immunoglobulin G (IgG) and goat anti-rat IgG marked with horseradish peroxidase as an example. In driving the reagent solutions, the average velocity of droplets is up to 5-7 mm/s on the ELISA chip, and a chip-scaled immunoassay can be finished within 20 min by using colorimetric detection, with the volume of the sample and reagents of only 0.5-1 mu l. (C) Koninklijke Brill NV, Leiden, 2011