High-throughput microarray-based enzyme-linked immunosorbent assay (ELISA).

Abstract

A new generation biochip is described as capable of supporting high-throughput (HT), multiplexed enzyme-linked immunosorbent assays (ELISAs). These biochips consist of an optically flat, glass plate containing 96 wells formed by an enclosing hydrophobic Teflon mask. The footprint dimensions of each well and the plate precisely match those of a standard microplate. Each well contains four identical 36-element arrays (144 elements per well) comprising 8 different antigens and a marker protein. Arrays are formed by a custom, continuous flow, capillary-based print head attached to a precise, high-speed, X-Y-Z robot. The array printing capacity of a single robot exceeds 20,000 arrays per day. Arrays are quantitatively imaged using a custom, high-resolution, scanning charge-coupled device (CCD) detector with an imaging throughout of 96 arrays every 30 s. Using this new process, arrayed antigens were individually and collectively detected using standard ELISA techniques. Experiments demonstrate that specific multiplex detection of protein antigens arrayed on a glass substrate is feasible. Because of the open microarray architecture, the 96-well microarray format is compatible with automated robotic systems and supports a low-cost, highly parallel assay format. Future applications of this new high-throughput screening (HTS) format include direct cellular protein expression profiling, multiplexed assays for detection of infectious agents and cancer diagnostics.