Focusing analytes from 50 μL into 500 pL: On-chip focusing from large sample volumes using isotachophoresis

Abstract

The use of on-chip isotachophoresis assays for diagnostic applications is often limited by the small volumes of standard microfluidic channels. Overcoming this limitation is particularly important for detection of ‘discrete’ biological targets (such as bacteria) at low concentrations, where the volume of processed liquid in a standard microchannel might not contain any targets. We present a novel microfluidic chip that enables ITP focusing of target analytes from initial sample volumes of 50 μL into a concentrated zone with a volume of 500 pL, corresponding to a 100,000-fold increase in mean concentration, and a 300,000-fold increase in peak concentration. We present design considerations for limiting sample dispersion in such large-volume focusing (LVF) chips and discuss the trade-off between assay time and Joule heating, which ultimately governs the scalability of LVF designs. Finally, we demonstrate a 100-fold improvement of ITP focusing performance in the LVF chip as compared to conventional microchannels, and apply this enhancement to achieve highly sensitive detection of both molecular targets (DNA, down to 10 fM) and whole bacteria (down to 100 cfu/mL).

Practical sample pretreatment techniques coupled with capillary electrophoresis for real samples in complex matrices

“…Later developments involve the use of microfluidic devices, as pioneered by Santiago et al [90]. Thus, ITP is a promising sample pretreatment technique for nucleic acids providing lower losses of trace analytes and EFs of several orders of magnitude [91,92,93,94], in contrast to current techniques such as SPE. Applications include DNA purification, extraction, fractionation, and extraction from single cells and/or hybridization.…”

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips (2016–2018)

“…Kooten et al . reported a large‐volume focusing (LVF)‐chip design whereby at initial, a larger sample volume can be used (50 μL) and preconcentrated into a concentrated zone with a volume of 500 pL using ITP. The LVF chip has a wide region tapering down to 100‐fold narrower channel together with a geometrical feature designed to reduce dispersion arising from the non‐uniform entry of the ITP interface into the narrow region.…”

Recent progress in analytical capillary isotachophoresis

“…Van Kooten and co‐workers decribed a novel design of a large‐volume microfluidic chip for ITP, with a wide region tapering down to a 100‐fold narrower channel. Its geometry is designed to reduce dispersion accompanying the entry of the ITP stack into the narrow region.…”