Abstract
In-line preconcentration techniques are used to improve the sensitivity of microfluidic DNA analysis platforms. The most common methods are electrokinetic and require an externally applied electric field. Here we describe a microfluidic DNA preconcentration technique that does not require an external field. Instead, pressure-driven flow from a fluid-filled microcapillary into a lower ionic strength DNA sample reservoir induces spontaneous DNA migration against the direction of flow. This migratory phenomenon that we call Molecular Rheotaxis initiates in seconds and results in a concentrated DNA bolus at the capillary orifice. We demonstrate the ease with which this concentration method can be integrated into a microfluidic total analysis system composed of in-line DNA preconcentration, size separation, and single-molecule detection. Paired experimental and numerical simulation results are used to delineate the parameters required to induce Molecular Rheotaxis, elucidate the underlying mechanism, and optimize conditions to achieve DNA concentration factors exceeding 10,000 fold.
Highlights
In-line preconcentration techniques are used to improve the sensitivity of microfluidic DNA analysis platforms
Evaporation and solid-phase extraction-based concentration methods have both been demonstrated in microfluidic devices[9,10,11,12], but electrokinetic methods are the most common and have demonstrated the highest nucleic acid concentration factors[13, 14]
Despite the popularity of these electrokinetic methods, there remain several challenges to their implementation in microfluidic devices: namely the requirements of embedded electrodes, external power supplies, microchannel wall conditioning and treatment to reduce electroosmotic flow[18, 19], and design considerations that avoid electric-fieldinduced DNA aggregation and degradation[20]. It was during the development of an electrokinetic concentration method for capillary sampling that we observed that certain conditions spontaneously triggered dilute DNA to collect into a highly concentrated bolus even without an applied electric field
Summary
In-line preconcentration techniques are used to improve the sensitivity of microfluidic DNA analysis platforms. Since SML-FSHS is performed using only a buffer-filled microchannel without sieving matrices or drag-tag conjugates to modulate mobility, wall coatings, or applied electric fields[27, 28], interfacing with MRT provides a truly electrode-free platform for highly sensitive and quantitative DNA preconcentration, size separation, and single-molecule detection. Both experimental and simulation results indicate that the technique is capable of effective DNA preconcentration with minimal size-dependent bias, demonstrating future applicability to a wide range of nucleic acid samples, assays, and applications. Using optimized conditions with the fully integrated preconcentration and separation platform, we demonstrate concentration factors exceeding 10,000 fold with HindIII digested λ DNA from a starting concentration as low as 150 aM
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call