Nanofluidic Biomolecular Preconcentration and Concentration-Enhanced Assays

Sample preparation is one of the bottlenecks in molecular detection and analysis. During the past decades, significant progress has been made both in binding assays (immunoassays) and mass spectrometry (MS). However, issues related to limited sample capacity and low abundance target create challenges in fully utilizing the power of these new analysis platform. In general, only high-abundance species of a given sample could be detected, while reliable analysis of low-abundance targets is still challenging. To address these problems, our group has sought ways to efficiently concentrate biomolecules in order to enhance the detection sensitivity for both large and small sample volumes. The nanofluidic electrokinetic concentration devices serve as;

(1) Ideal world-to-microchip coupling system: It can collect biomolecules from ~µL fluidic samples (addressable by pipettes) and concentrate them into ~nL plug (addressable by microfluidics).
(2) Generic sensitivity enhancement scheme for many biochemical assays: Wide variety of biochemical assays can be enhanced simply by collecting the reactants and / or target molecules.

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 target-bound 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 fully-automated 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.