Dietrich Kohlheyer

Miniaturizing free-flow electrophoresis – a critical review

Free‐flow electrophoresis (FFE) separation methods have been developed and investigated for around 50u2005years and have been applied not only to many types of analytes for various biomedical applications, but also for the separation of inorganic and organic substances. Its continuous sample preparation and mild separation conditions make it also interesting for online monitoring and detection applications. Since 1994 several microfluidic, miniaturized FFE devices were developed and experimentally characterized. In contrast to their large‐scale counterparts microfluidic FFE (μ‐FFE) devices offer new possibilities due to the very rapid separations within several seconds or below and the requirement for sample volumes in the microliter range. Eventually, these μ‐FFE systems might find application in so‐called lab‐on‐a‐chip devices for real‐time monitoring and separation applications. This review gives detailed information on the results so far published on μ‐FFE chips, comprising its four main modes, namely free‐flow zone electrophoresis (FFZE), free‐flow IEF (FFIEF), free‐flow ITP (FFITP), and free‐flow field‐step electrophoresis (FFFSE). The principles of the different FFE modes and the basic underlying theory are given and discussed with special emphasis on miniaturization. Different designs as well as fabrication methods and applied materials are discussed and evaluated. Furthermore, the separation results shown indicate that similar separation quality with respect to conventional FFE systems, as defined by the resolution and peak capacity, can be achieved with μ‐FFE separations when applying much lower electrical voltages. Furthermore, innovations still occur and several approaches for hyphenated, more integrated systems have been proposed so far, some of which are discussed here. This review is intended as an introduction and early compendium for research and development within this field.

Free-flow zone electrophoresis and isoelectric focusing using a microfabricated glass device with ion permeable membranes

This paper describes a microfabricated free-flow electrophoresis device with integrated ion permeable membranes. In order to obtain continuous lanes of separated components an electrical field is applied perpendicular to the sample flow direction. This sample stream is sandwiched between two sheath flow streams, by hydrodynamic focusing. The separation chamber has two open side beds with inserted electrodes to allow ventilation of gas generated during electrolysis. To hydrodynamically isolate the separation compartment from the side electrodes, a photo-polymerizable monomer solution is exposed to UV light through a slit mask for in situ membrane formation. These so-called salt-bridges resist the pressure driven fluid, but allow ion transport to enable electrical connection. In earlier devices the same was achieved by using open side channel arrays. However, only a small fraction of the applied voltage was effectively utilized across the separation chamber during free-flow electrophoresis and free-flow isoelectric focusing. Furthermore, the spreading of the carrier ampholytes into the side channels resulted in a very restricted pH gradient inside the separation chamber. The chip presented here allows at least 10 times more efficient use of the applied potential and a nearly linear pH gradient from pH 3 to 10 during free-flow isoelectric focusing could be established. Furthermore, the application of hydrodynamic focusing in combination with free-flow electrophoresis can be used for guiding the separated components to specific chip outlets. As a demonstration, several standard fluorescent markers were separated and focused by free-flow zone electrophoresis and by free-flow isoelectric focusing employing a transversal voltage of up to 150 V across the separation chamber.

A prefilled, ready-to-use electrophoresis based lab-on-a-chip device for monitoring lithium in blood

We present the Medimate Multireader, the first point-of-care lab on a chip device that is based on capillary electrophoresis. It employs disposable pre-filled microfluidic chips with closed electrode reservoirs and a single sample opening. Several technological innovations allow operation with closed reservoirs, which is essential for reliable point-of-care operation. The chips are inserted into a hand-held analyzer. In the present application, the device is used to measure the lithium concentration in blood. Lithium is quantified by conductivity detection after separation from other blood ions. Measurements in patients show good accuracy and precision, and there is no difference between the results obtained by skilled and non-skilled operators. This point-of-care device shows great promise as a platform for the determination of ionic substances in diagnostics or environmental analysis.

Synchronized, continuous-flow zone electrophoresis.

A new method for performing continuous electrophoretic separation of complex mixtures in microscale devices is proposed. Unlike in free-flow electrophoresis devices, no mechanical pumping is required--both fluid transport and separation are driven electrokinetically. This gives the method great potential for on-a-chip integration in multistep analytical systems. The method enables us to collect fractionated sample and tenfold purification is possible. The model of the operation is presented and a detailed description of the optimal conditions for performing purification is given. The chip devices with 10-microm-deep separation chamber of 1.5 mm x 4 mm in size were fabricated in glass. A standard microchip electrophoresis setup was used. Continuous separation of rhodamine B, rhodamine 6G, and fluorescein was accomplished. Purification was demonstrated on a mixture containing rhodamine B and fluorescein, and the recovery of both fractions was achieved. The results show the feasibility of the method.

Reaction and diffusion dynamics in a microfluidic format

A novel microfluidic structure based on the electroosmotic guiding of reagent streams is presented that can be used as a fully adjustable diffusion based microreactor. The position and the width of two aqueous reactant streams entering a laminar-flow chamber can be controlled individually by changing the flow ratio of three parallel guiding streams containing buffer only. To control the intensity of product formation, the overlapping area between the diffusion regions of the two different reagent streams can be adjusted. This article describes the fabrication and experimental characterization of the device.