On-Chip Millionfold Sample Stacking Using Transient Isotachophoresis

Abstract

We present a simple and robust isotachophoresis (ITP) method that can be integrated with microchip-based capillary electrophoresis (CE) devices to achieve millionfold sample stacking. We performed an experimental parametric study to show the effects of initial sample ion concentration, leading ion concentration, and trailing ion concentration on ITP stacking. We also discuss the usefulness and limitations of a simple one-dimensional nondispersive model and a scaling analysis for dispersion rate. We found that a single-column ITP configuration together with electroosmotic flow suppression and high leading ion concentration provide high-performance ITP and can be integrated readily with CE separation. We demonstrated detection of trace of 100 fM Alexa Fluor 488 (signal-to-noise ratio of 11) with a concentration increase of a factor of 2 x 10(6). Application of our ITP/CE protocol to the stacking and separation of negatively charged fluorescent tracers (Alexa Fluor 488 and bodipy) resulted in a concentration increase of 6.4 x 10(4) and a signal increase of 4.5 x 10(5). The ITP/CE protocol can be performed with a standard microchannel cross design or simple flow control. The method can be implemented with available off-the-shelf chip systems using off-the-shelf voltage control systems and buffer chemistries.

Bacterial RNA Extraction and Purification from Whole Human Blood Using Isotachophoresis

“…ITP is a highly sensitive, 28,29 robust 30,31 sample preparation method that can, under ideal conditions, provide up to 1,000,000-fold preconcentration. 32 We have demonstrated successful ITP extraction of small RNA from cell culture lysate, 33 micro-RNA from total RNA, 34,35 genomic DNA (gDNA) 36 and pathogenic DNA (malaria) 37 from whole blood lysate, and rRNA from bacteria in urine lysate. 38 However, no previous nucleic acid extraction protocols using ITP have been designed for adequate RNase control and RNA integrity.…”

Microsystem for Isolation of Fetal DNA from Maternal Plasma by Preparative Size Separation

“…A drawback of these techniques is that they allow only relatively small sample volumes (a few microliters) to be used, and the target molecules are not always nucleic acids. Transient ITP offers the advantage of size separation after preconcentration (34 ), which facilitates the concentration of a DNA sample before separation (35 ); however, transient ITP suffers from the drawback of requiring long channels that exceed the chip dimensions for samples Ͼ10 L. Other approaches for preconcentration have exploited filter structures or membranes (36,37 ) and, recently, electrokinetic trapping (EKT) at polyethylene terephthalate (PET) membranes for stacking larger volumes of about 80 L (38 ).…”

Integrated microfluidic preconcentrator and immunobiosensor

“…A number of preconcentration techniques have been developed that can achieve high concentration factors in small time durations. Some examples include surfacebinding techniques like solid-phase extraction (Jemere et al 2002;Ramsey and Collins 2005;Yu et al 2001) and electrokinetic manipulation techniques like isoelectric focusing (Li et al 2003;Tan et al 2002;Cabrera and Yager 2001), field-amplified sample stacking (Jung et al 2003;Lichtenberg et al 2001), isotachophoresis (Jung et al 2006;Wainright et al 2002;Wang et al 2009), and dielectrophoresis (Lapizco-Encinas et al 2005;Moncada-Hernandez and Lapizco-Encinas 2010). However, the limitations of these techniques are that they either involve buffer handling challenges or fabrication complexities making them difficult to integrate with lab-on-chip systems.…”