Increasing the Sensitivity of ELISA using Multiplexed Electrokinetic Concentrator

We developed a novel method to increase the sensitivity of standard Enzyme-Linked Immunosorbent Assay (ELISA) using a multiplexed electrokinetic concentration chip. The poly(dimethylsiloxane) (PDMS) molecular concentrator1 was used to trap and collect charged fluorescent product of targetbound enzyme turnover reaction of ELISA that occurred in a standard 96 well plate. Detection sensitivities of both Prostate Specific Antigen (PSA) and CA 19-9 (a human pancreatic and gastrointestinal cancer marker) ELISAs in serum are enhanced ~100 fold with a low CV of <17%. We also integrated this method with an on-chip bead-based ELISA that lends itself toward a fullyautomated on-chip diagnostic device. Detection sensitivity of microfluidic bead-based CA 19-9 ELISA in serum is enhanced ~65 fold compared to the results without electrokinetic accumulation step. This chip can be directly applied to enhance the readout sensitivity of a wide range of existing ELISA kits at concentrations below the current detection limit.