Continuous microfluidic DNA extraction using phase-transfer magnetophoresis

Abstract

This paper reports a novel microfluidic-chip based platform using "phase-transfer magnetophoresis" enabling continuous biomolecule processing. As an example we demonstrate for the first time continuous DNA extraction from cell lysate on a microfluidic chip. After mixing bacterial Escherichia coli culture with superparamagnetic bead suspension, lysis and binding buffers, DNA is released from cells and captured by the beads. These DNA carrying beads are continuously transported across the interfaces between co-flowing laminar streams of sample mixture, washing and elution buffer. Bead actuation is achieved by applying a time-varying magnetic field generated by a rotating permanent magnet. Flagella-like chains of magnetic beads are formed and transported along the microfluidic channels by an interplay of fluid drag and periodic magnetic entrapment. The turnover time for DNA extraction was approximately 2 minutes with a sample flow rate of 0.75 µl s(-1) and an eluate flow rate of 0.35 µl s(-1). DNA recovery was 147% (on average) compared to bead based batch-wise extraction in reference tubes within a dilution series experiment over 7 orders of magnitude. The novel platform is suggested for automation of various magnetic bead based applications that require continuous sample processing, e.g. continuous DNA extraction for flow-through PCR, capture and analysis of cells and continuous immunoassays. Potential applications are seen in the field of biological safety monitoring, bioprocess control, environmental monitoring, or epidemiological studies such as monitoring the load of antibiotic resistant bacteria in waste water from hospitals.

Controlled counter-flow motion of magnetic bead chains rolling along microchannels

“…This was possible even for a counter-flow arrangement with a mean liquid velocity of up to -8.0 mm s -1 . The use of this method within a real world biological assay implementation such as continuous DNA extraction has been shown in (Karle et al 2010). Furthermore, the walking bead chains enable a wide range of other Fig.…”

On-line cell lysis of bacteria and its spores using a microfluidic biochip

“…The final microfluidic channel structure of the lysis chamber can be determined with 5 ml of internal volume, where the sample and lysis buffers are mixed and lysed in one step. Despite the fact that the purification process of DNA by washing and elution steps is not integrated in the biochip, successful cell/spores lysis was achieved on a microfluidic chip [30]. Continuous flow lysis protocol allows the experimenter to lyse bacteria in a sample with a volume of 20 ml for 20 minutes in the presence of 40 kPa overpressure.…”

Centrifugal LabTube platform for fully automated DNA purification and LAMP amplification based on an integrated, low-cost heating system

“…T he following primers and probes (Karle et al 2010) were used at 200nM in conjunction with the Quantifast master mix and Rox -dye (Qiagen, Cat.no. 204352…”