EP1188048A1

EP1188048A1 - Miniaturized analytical system with a device for withdrawing substances

Info

Publication number: EP1188048A1
Application number: EP00940331A
Authority: EP
Inventors: Friedhelm Eisenbeiss; Bernd Stanislawski; Thomas Greve; Roland HERGENRÖDER; Günther Weber; Benedikt Grass; Andreas Neyer; Matthias JÖHNCK
Original assignee: Merck Patent GmbH
Current assignee: IP VERWERTUNGS GmbH
Priority date: 1999-06-16
Filing date: 2000-06-06
Publication date: 2002-03-20
Also published as: AU5530200A; DE19927535B4; DE19927535A1; JP2003502636A; WO2000077508A1

Abstract

The invention relates to a device for withdrawing defined components of samples from the separation channel and transporting them to another channel after a preparative or analytical liquid phase separation in flat, miniaturized analytical systems. The withdrawal device can be directly integrated into the analytical system by positioning detectors, for example for measuring the conductivity, any place in the analytical system.

Description

Miniaturized analysis system with device for discharging substances

The invention relates to a device for discharging defined fractions of samples after a preparative or analytical

Liquid phase separation in planar, miniaturized analysis systems from the separation channel into another channel or into another analytical device.

Planar, miniaturized analysis systems consist of components with integrated channels in which the transport and / or separation of dissolved analytes takes place, for example, by means of capillary electrophoresis or isotachophoresis. Such a channel system can have Y-shaped branches and / or X-shaped crossings (see Figure 1). The angles between the channels are freely selectable.

An X-shaped arrangement of the channels is usually used for introducing sample material, and a Y-branching for discharging. For this purpose, the sample components are transported electrokinetically by applying a voltage to the ends of the channels. In the case of Y-branched channels, for example, the electrokinetic transport can be redirected if the electrical potentials are switched from one channel to the other branching channel. In this way, a fraction of a sample can be discharged through the branching channel.

Such discharge processes are controlled either in a time-defined manner or actively controlled in the case of a previous passage analysis. Precise knowledge of the electropherogram and exact reproducibility of the separation process are required for time-controlled discharge. The analyte to be isolated can therefore only be obtained from a known sample after its experimental determination Separation time can be removed. This method is particularly unsuitable for capillary electrophoresis, since the addition of surface-active substances leads to a change in the zeta potential on the channel walls. This causes a modulation of the electroosmotic force and thus also a modulation of the temporal pattern of the

Electropherogram. For this reason, time-controlled rejection is usually only carried out for checking or confirmation.

Removal of the analytes after a direct analysis is much more precise. By F. von Heeren et al. (Anal. Chem 68 (13) (1996), 2044-2053) describes the actively controlled removal of sodium fluorescein. The position of the fluorescein during the separation process can be continuously monitored by an observer using a suitable optical device. As soon as the fluorescein band is at the discharge point, a switching process is triggered manually, which leads to the discharge. However, even when this system is automated, only colored or fluorescent substances could be detected and specifically discharged. This means a severe limitation.

Other detector systems for ejection devices have so far not been able to be integrated directly into miniaturized, planar analysis systems, so that substances are usually detected and separated after they have left the analysis system.

The object of the present invention is therefore to provide a device for discharging substances for miniaturized planar analysis systems which is integrated directly into the analysis systems and can be actively controlled. The device for discharging should preferably be combinable with detection devices which are based on different principles; thus the removal of analytes would be very versatile. For planar devices for electrophoretic separation processes, an arrangement comprising at least three transport electrodes, a detection device and a switching device has been found, which makes it possible to selectively discharge fractions during the separation process. In preferred embodiments, the detection device is designed as an electrical conductivity, impedance or potential measuring device.

The invention therefore relates to a device for discharging fractions of a sample for planar microstructured analysis systems, which essentially consist of a channel system with at least one Y branch, at least three transport electrodes and at least one detection device in front of said branch point of the channel system and an electrical switching device.

A preferred embodiment of the invention is a device for discharging, in which the detection device is an electrochemical detector.

The present invention furthermore relates to the use of a device according to the invention in a planar microstructured analysis system.

Figure 1 shows an example of an X crossing (X) and a Y branch (Y) of a channel system according to the prior art.

Figure 2 illustrates the principle of discharge using the device according to the invention. Figures 3 and 4 schematically show analysis systems in which an inventive device for discharging substances is integrated.

Analysis systems in which an inventive device for

Discharge of substances that can be integrated are planar microstructured systems that serve to separate substances. Such predominantly two-dimensional analysis systems, due to their small size and simple manufacture, offer many advantages over macro-scope analysis systems. The analysis systems can include additional analysis devices or devices for micro-preparative derivatization. Due to the possibility of integrating detectors or a device according to the invention for discharging directly into these systems during the manufacture of analysis systems, substances can be analyzed and separated in the analysis system already during or after the separation. For example in analysis systems in which substances are not only separated and analyzed, but also other e.g. Are subjected to derivatization steps, more than one device for discharging according to the invention can also be integrated.

The discharge device according to the invention consists of a detector system which analyzes a channel segment directly in front of the discharge channel, and a branched channel system with corresponding transport electrodes. As soon as the detector indicates that the desired analyte is just before the branch, the transport electrodes are switched at the ends of the channels. The further transport no longer takes place along the separation channel but in the branching discharge channel. This process ends when the detector indicates that the analyte band has passed the ejection point. In this way, defined parts of a sample can be precisely separated from the rest of the sample. The process of discharging therefore comprises the following steps:

• The spatially separated sample is transported to a branching point by appropriate electrical voltage. • A detector, which sits close to the branching point, measures the components flowing past or a marker substance, which marks the beginning and the end of an area to be discharged.

• As soon as the desired component is detected, the voltages are switched so that the flow in the branches

Channel is directed.

• After the desired component has passed the detector, the voltage is switched back.

In this way, part of the sample is now spatially separated from the rest of the sample.

The entire process, in particular the switching of the potentials between the electrodes, is preferably controlled by means of an electronic switching device. Such devices and their use are known to the person skilled in the art. The separated substances can then be subjected to further steps, such as separate analyzes, derivatizations, etc., within the analysis system. Furthermore, they can also be removed from the channel system of the analysis system. For this purpose, the duct system is provided with additional outputs. These outputs are preferably located in the discharge channels and are closed off by means of a fluid connection, such as a tightly closing pump or pumps and valves. Typically, a capillary directly follows, via which the separated fraction can be transferred to other containers or devices outside of the analysis system. Accordingly, if there is a separated fraction in a discharge channel, it can 0/77508. g. PCT / EPO0 / 052O5

they are removed hydromechanically, for example electroosmotically or by means of micropumps, from an outlet from the analysis system.

The removed fraction can be bothersome

Act a component that is to be separated so that the rest of the sample can be examined further, as well as a fraction, i.e. a part of the sample which is to be subjected to further analysis or derivatization steps separately from the rest of the sample. The sample components to be analyzed and separated in this way can be ionically dissolved, emulsified, suspended, colloidal or biologically cellular in predominantly aqueous solution.

The device according to the invention is based on a detector located on the two-dimensional analysis system. Since miniaturized analysis systems can be structured very differently depending on their specific application, the detector must be able to be positioned anywhere in the analysis system. Both detectors can be used for the device according to the invention, the most important parts of which can be integrated, such as

- conductivity detectors (capacitive, inductive, ohmic measurement),

- electrochemical detectors (e.g. amperometry, ISFET),

- electrical temperature measurement, but also external detectors in which only parts, e.g. Lenses or fiber optics can be integrated into the analysis system, such as

- optical detectors (e.g. refractive index, temperature, absorption, fluorescence, Raman, luminescence)

- NMR

- radioactive labeling - magnetic labeling

According to the invention, conductivity detectors and optical detectors are preferably used as the detector device. For optical detectors For example, a recording device for optical fiber optics can be integrated.

For analysis systems in which substances are separated electrophoretically, the requirement for a universal detection method is particularly well met by electrical detection methods such as conductivity measurement. The analytes are characterized by their special electrical conductivity. A given substance always generates the same relative conductivity in a given electrolyte system. This applies to successive measurements in a miniaturized

Analysis systems and also for measurements that are carried out in several miniaturized analysis systems of one type.

An electrical conductivity measurement is therefore preferably used in the device according to the invention, which measures the electrical current or the electrical voltage drop in the case of directly contacting electrodes or, in the case of galvanically decoupled electrodes, is carried out by measuring the dielectric resistance.

To integrate a conductivity detector in a two-dimensional analysis system, conductivity electrodes must be integrated at any point in the system, especially shortly before branches along the channel system. This is only possible through a special type of construction of such a system. On the one hand, the channel system must be sealed gas and liquid-tight, on the other hand, it must be ensured that chemically inert electrodes can be attached precisely and reproducibly at the desired positions. The above-mentioned requirements can only be met by special techniques for the production of two-dimensional analysis systems.

Microfluids or microstructured analysis systems usually consist of a flow unit, which is at least the channel system and optional Has recesses for the integration of peripheral devices, and peripheral devices, such as detectors, fluid connections, storage vessels, reaction chambers, pumps, control devices etc., which can be integrated into the flow unit or connected to it. According to the invention, flow units for microfluidic analysis systems with measuring and control devices for electrical conductivity are systems in which microchannel structures are produced by joining at least two components, such as substrate and cover, which can be closed in a liquid-tight and / or gas-tight manner.

The systems according to the invention typically consist of at least two components, a cover which is provided with the electrodes and a microstructured substrate. After the components have been produced, they are joined using a special bonding process. In this way, it is possible to integrate the discharge device according to the invention in planar analysis systems.

The components of the flow unit of the analysis systems preferably consist of commercially available thermoplastics, such as PMMA (polymethyl methacrylate), PC (polycarbonate), polystyrene or PMP (polymethylpentene), cycloolefinic copolymers or thermosetting plastics, such as epoxy resins. All components of a system preferably consist of the same material.

The components can be produced by methods known to the person skilled in the art. Components that contain microstructures can be produced, for example, by established processes such as hot stamping, injection molding or reaction molding. Components which can be reproduced using known techniques for mass production are particularly preferably used. Microstructured components can have channel structures with cross-sectional areas between 10 and 250,000 μm ² . For the According to the discharge device according to the invention, in addition to areas for sample application and a separation channel, the channel system must have at least one X or Y branch originating from a separation channel. In order to integrate several ejection devices, further branches can be introduced at any point in the channel system.

The electrodes which are required for the discharge device according to the invention are transport electrodes which are located at the ends of the branched channels and enable the potential to be switched between the two channels, and detection electrodes which are preferably between 40 mm and 0.1 μm before the branch are positioned.

To integrate the electrodes into the analysis system or the device according to the invention, the electrodes are preferably attached to a component of the system, the cover. To do this, they must have sufficient adhesive strength on the plastic component. This is important for the assembly of the individual components as well as for the later use of the analysis systems. If, for example, If adhesives are used, the adhesive must not detach the electrode from the plastic surface. Furthermore, the electrodes should be made of chemically inert materials such as e.g. Precious metals (platinum, gold) exist.

Plastic surfaces are typically metallized by electrochemical deposition of metals from metal salt solutions.

For this purpose, it is common practice to first pretreat the plastic surface chemically or mechanically in a multi-stage process, to apply a discontinuous primer and finally to carry out the electrochemical deposition. Descriptions of these metallization techniques can be found, for example, in US Pat. No. 4,590,115, EP 0 414 097, EP 0 417 037 and in Wolf and Gieseke (DG Wolf, H. Gieseke, "New Process for All-Over and Partial Metallization of Art Stoffen, "Galvanotechnik 84, 2218-2226, 1993). Common to the wet chemical processes is that relatively complex pretreatment processes are necessary in order to achieve sufficient adhesive strengths.

DE 196 02 659 describes the adherent application of copper to multiphase polymer mixtures by means of vapor deposition or sputtering. The composition of the polymer mixtures is mentioned as the cause of the good adhesion. Accordingly, the mixtures must contain polyarylene sulfides, polyimides or an aromatic polyester.

The influence of plasma pretreatments to achieve better adhesive properties of metals on plastic surfaces is summarized by Friedrich (J. Friedrich, "Plasma treatment of polymers", kleben & dichten 41, 28-33, 1997) using the example of various commercially available thermoplastics.

The electrode structures on the plastic components are particularly preferably produced by means of a two-layer technique. For this purpose, an adhesion-promoting layer made of chromium oxide is first created. In contrast to precious metals, chromium oxide shows excellent adhesive properties on plastic surfaces. In addition, unlike elemental chromium and other transition metals, chromium oxide is much more resistant to redox processes. The noble metal, such as platinum or its alloys or gold, is then applied to the chromium oxide adhesive layer.

The selective application of chromium oxide and the noble metal layer to be deposited thereon is preferably carried out on plastic substrates using the lift-off method or by means of the shadow mask technique or the structuring of metallic layers initially applied over the entire surface. These process technologies are standard processes in microstructure technology. in the In the following, the work steps required for the two-layer technique for the mentioned processes are briefly described.

Lift-off process: The plastic component to be selectively metallized is coated with a photoresist. This photoresist must not or only slightly dissolve the plastic part to be metallized. For PMMA e.g. a photoresist from Allresist, Berlin (AR 5300/8) has proven to be suitable. After exposure and development of the structures to be metallized, the metallic layers are applied in a sputtering system. The chromium oxide layer is applied during the sputtering process by introducing oxygen into the typically used argon plasma of the sputtering system. A conventional chrome target is used as the sputtering target. Typical chromium oxide layer thicknesses are 10-50 nm. Alternatively, a chromium oxide target can be used directly. The sputtering of platinum or its alloys or of gold is carried out immediately afterwards under standard conditions, i.e. in argon plasma. It has also proven to be advantageous for the adhesive strength of the chromium oxide layer to have the plastic sputtered back in an oxygen / argon (approx. 5 vol% / 95 vol%) plasma before the chromium oxide is sputtered. In the actual lift-off process, the still existing photoresist and with it the metal layer on the lacquer are detached from the plastic component in a developer from Allresist (AR 300-26).

Shadow mask technique: The plastic part to be selectively metallized is covered with a so-called shadow mask. This has cutouts in the areas to be metallized. Through this, the metal layers are tracked down in analogy to the lift-off process!

Structuring of flat metallic layers: A metal layer is first applied to the entire surface of a plastic part that is to be selectively metallized, analogously to the sputtering process already described. This is structured in subsequent process steps, either by selective ablation using, for example, laser ablation (gold and platinum) or, for example, by selective wet-chemical etching. For structuring by means of wet chemical etching, a photoresist (Hoechst AG, Germany; AZ 5214) is first applied, exposed and developed on the metal layer. Gold is then stripped off in cyanide solution in the exposed areas. The electrically non-conductive chromium oxide layer remains. Finally, the remaining photoresist is removed with a developer (eg AR 300-26, Allresist, Berlin).

The adhesive strength of electrodes produced with chromium as well as with chromium oxide as an adhesive layer using sputter technology was checked with the aid of tear tests. The adhesive strength of the chrome oxide layers is significantly greater. Even in the case of ultrasound treatment in alkaline solution, the metal layers which have been produced using chromium oxide as the adhesive layer are significantly more stable compared to metal layers which have been produced using chromium as the adhesive layer.

After production and preparation of the individual components, these are assembled using the method according to the invention. A component, the substrate, is preferably microstructured and provided with bores or cutouts on the back for filling the channels and / or contacting the electrodes. Furthermore, the use of a so-called sealing lip, i.e. an elevation completely surrounding the channel structures on the substrates with heights between typically 0.5 to 5 μm, has proven to be very advantageous with regard to the bonding process. The other component, the lid, is used for covering and is e.g. with electrodes in electrophoretic analysis systems. In this case, the lid is referred to as an electrode lid according to the invention. For certain applications, the

Analysis systems require a functionalization of the components that deviates from this preferred arrangement. In this case, you can For example, more than two components, for example two lids and a substrate, etc., are joined together to produce superimposed channel structures, or further functionalities, such as detection systems, reaction chambers, etc., are integrated into the components. According to the invention, all parts of the flow unit of the analysis system that are joined together by means of a bonding method are referred to as components. They can be microstructured, provided with electrodes or have other functionalities. A subdivision of the components into substrates and cover or also electrode cover, if the corresponding component is provided with electrodes, only serves the closer

Description of the embodiment of the special components and does not constitute a restriction with regard to further properties of the components, such as microstructuring etc., or their combination with one another.

The assembly of the components takes place according to the invention with high precision. The adhesive must not run into the channels and cover their surface, as this can change the surface properties of the channels. It has been found that this leads, for example, to increased adhesion of analytes, e.g. Proteins, leads to the channel areas that are wetted with adhesive. This in turn affects the separation quality of the analysis systems. Likewise, gluing the electrodes with adhesive affects their functionality.

It is also of great importance that the volume of the channels is not changed, as would be the case, for example, due to the uncontrolled inflow of adhesive. According to the invention, the channel for improving the detection sensitivity is preferably narrowed in the vicinity of the detection electrodes. It is important in these areas that no glue gets into the channel.

To assemble the components, it is preferred according to the invention firstly to the microstructured component at the locations where none 00/77508. 14. PCT / EP0O / O52O5

Structuring is present, an adhesive is applied. The layer thickness is between 0.5 and 10 μm, preferably between 3 and 8 μm. Typically, the application takes place by means of a flat roller application known from printing technology.

In a preferred embodiment, a thin film of adhesive is applied to a second unstructured roller, which is coated with a polymer, by means of a structured metallic anilox roller which holds a defined volume of adhesive. This in turn is applied directly to the structured substrate in such a way that there is preferably an adhesive thickness between 3 and 8 μm on the unstructured surface of the substrate. Depending on the plastic used (substrate material), the transfer between the plastic roller and the substrate is influenced by a possible increase in the viscosity of the adhesive (prepolymerization). An important advantage of this method is that the substrate does not have to be positioned relative to the roller applying the adhesive, and nevertheless adhesive is only applied in the non-structured areas of the substrate. If too much adhesive is applied, adhesive will flow into the channel when the lid and substrate are pressed together. If insufficient adhesive has been applied in some areas, the channel structure will leak. This connection method requires a flatness of the components of preferably less than approx. 5 // m / cm component length.

The adhesive used must not or only very slightly dissolve the surface of the components so that the electrodes are not detached or interrupted by the adhesive during the bonding process. The product NOA 72, thiol acrylate from Norland, New Brunswick, NJ, USA is therefore preferably used as the adhesive. This adhesive is cured photochemically. However, other types of adhesives, such as, for example, thermosetting adhesives, which meet the above-mentioned requirements can also be used for the method. After the adhesive has been applied, the second component with the thin-film electrodes is suitably positioned and pressed onto the substrate, for example on an exposure machine. For this purpose, the substrate with the applied adhesive is preferably fixed in the exposure machine in the position otherwise provided for silicon wafers. The use of strong glass plates as the pressing surface is preferred since the positioning and the photochemical curing of the adhesive can be carried out directly by irradiation with an Hg lamp (emission wavelength 366 nm). The electrode cover is fixed in the position provided for the exposure mask by holding it with a vacuum device milled into a glass plate. Since both the electrode cover and the glass plate used to hold the cover are transparent, the cover can be adjusted with respect to the substrate through this arrangement. If the cover extends beyond the substrate, it can also be held mechanically.

The positioning of the lid on the substrate can typically take place in addition to an optical mechanical adjustment with the aid of optical adjustment marks, also passively mechanically with the aid of a snap-in device, optically mechanically without special adjustment marks or electrically mechanically with the aid of electrical marks (contacts).

The preferred optical metallic ones have been found

Adjustment marks applied to the lid in the same process step as the electrodes, i.e. preferably sputtered on, i.e. no additional effort is necessary. The corresponding counter structures on the substrate do not require any additional processing, since these are combined with the channel structures in one

Impression step can be introduced into the substrate. For the optical mechanical adjustment, at least one component must be made of a transparent one Plastic. With the aid of the alignment marks applied according to the invention, the two components are positioned with one another and pressed together with an accuracy of at least ± 10 μm, typically even ± 2 μm (for example the target position to the actual position of the detector electrode). The high positioning accuracy supports the realization of reproducible

Analysis results. Now the adhesive is polymerized with a UV lamp. After switching off the vacuum for the lid holder or loosening the mechanical fixation, the flow unit is removed from the exposure machine.

In another preferred embodiment, a component is provided with adhesive by means of a process known in printing technology (pad printing). For this purpose, the component provided with the electrodes is wetted with the adhesive on the areas which do not lie over a channel when the two components are assembled or which need to be electrically contacted. Microstructured components are wetted so that no adhesive gets into the channel structure or other recesses. The pad printing is a structured adhesive application. Adhesive is stored in a negative form of the substrate. This adhesive is absorbed in a structured manner by a typically silicone cushion and e.g. applied to the cover so that the areas that later form a wall of a fluidic channel are not wetted with adhesive. The component with the channel structures is then, as already described, suitably positioned and pressed onto its counterpart. The curing takes place as described above.

A structured adhesive application using spray techniques (e.g. microdrop process) or using the screen printing technique is also possible, provided that the lateral dissolution of the adhesive application is sufficient.

According to the invention, pressing the second component or pressing the components together means that the components are suitable be brought into contact with each other. In order to achieve a permanent connection of the components after curing, it is usually not necessary to exert a large force, that is to say to press the components very strongly together.

If the curing process of the adhesive is carried out outside of the adjustment device used for positioning the lid and substrate, the metallized lid and the substrate, after they have been adjusted to one another, can first be tacked by means of laser welding. The composite is then removed from the adjustment device and the adhesive used is cured in a separate exposure apparatus or an oven. This procedure means process acceleration and simplification, since curing no longer has to take place in the adjustment device.

Since the thermoplastic materials which are preferably used are largely transparent to laser light in the visible and near-infrared wavelength range, laser welding in this wavelength range requires an absorber layer for absorbing the optical power at the interface between the cover and the substrate. This

The absorber layer is applied simultaneously with the application of the power or detector electrodes. For example, when the electrodes are sputtered with noble metal, the electrode cover can additionally be sputtered with an Edei metal layer as an absorber layer at other points.

The welding of an electrode cover provided with 200 nm thick platinum electrodes, which therefore also includes additional platinum surfaces for absorbing the laser power, to a substrate (PMMA base material) is carried out using diode laser radiation (wavelength mixture of 808, 940 and 980 nm) with a power of 40 watts with a focus diameter of 1.6 mm. The platinum layer is destroyed during welding. Alternatively, the use of a substrate or cover filled with soot particles, for example, is also possible as an absorber. This latter procedure has the disadvantage that at least one channel wall is made of a different material. This also limits the possibilities of coupling optical power into or out of the channel for optical detection purposes.

The contacting of the transport and detection electrodes as well as the automatic control and the switching of the electric flow take place according to methods known to the person skilled in the art.

In this way, transport and detection electrodes can be integrated into the microstructured analysis systems in such a way that one or more discharge devices according to the invention are produced.

The integration of the discharge devices does not require any additional effort nor is the quality (stability, size, etc.) of the analysis systems influenced.

The device according to the invention thus represents an important additional functionality for planar microstructured analysis systems. For the first time, it enables the design of multifunctional microstructured analysis systems or flow units for analysis systems. The systems are not only able to separate samples, they can also separate, identify and select samples without the sample leaving the analysis system. This also opens up the possibility of only executing certain sample components after being removed from the analysis system, or of subjecting them to further derivatization or analysis steps in the system.

Closed microchannel structures can be achieved for the first time through the described manufacturing processes of the electrodes and bonding processes 0/77508. -jg. PCT / EP00 / 05205

are generated in which electrodes can be positioned anywhere within the channels. Structured components (substrates) can be provided in a liquid-tight and gas-tight manner, for example with electrode covers. By using mostly commercially available plastics and simple processing steps, such

Analysis systems can be produced inexpensively and in large numbers. Due to the special procedure for joining or bonding, the components are wetted with adhesive in such a way that after joining, no adhesive enters the interior of the duct system, i.e. gets into the channels, the walls or electrodes or other devices protruding into the channel system. This improves the separation quality and analysis sensitivity of the systems. They meet all the requirements that must be placed on a variably usable, precisely working analysis system: • They show high dimensional and volume stability of the channels.

• Due to the strength of the adhesive connections, they are inside the

Channels pressure stable.

• There is great variability in what can be used

Plastics. • Chemically inert materials can be used for components and electrodes.

• All four channel walls are preferably made of the same material.

• The electrodes can be positioned at any point on the channels with an accuracy of ± 10 μm.

• The contact surfaces of the electrodes are free from contamination

Adhesive.

• The electrodes can be easily connected.

• The systems show low internal resistance and allow potentially high current densities. Figure 2 illustrates the discharge of a substance using the device according to the invention. Three different stages of discharge are shown in pictures A, B and C. The schematic discharge device consists of a Y-branched channel system with the transport electrodes 1, 2 and 3 at the ends of the channels. The

Channel section between electrodes 1 and 2 serves as a separation channel, the channel branching off to electrode 3 is the discharge channel. Shortly before the discharge channel branches off, there is a detector electrode 4 in the separation channel. In Figure A, substances 5 and 6 to be separated migrate along the separation channel due to a potential between electrodes 1 and 2. Image B shows the moment in which the substance 5 being sought passes the detector electrode. The detected signal, for example the specific relative conductivity, causes the potential to be switched so that there is now a potential between electrodes 1 and 3. As a result, as shown in Figure C, substance 5 migrates into the discharge channel and is thus separated from substance 6, which is located in the separation channel. After substance 5 has passed the detector area and migrated into the discharge channel, the potential can be switched over again so that no further substances get into the discharge channel.

Figure 3 shows schematically an example of a miniaturized planar analysis system with an integrated device according to the invention for ejecting samples. The system contains a separation channel T1 with two transport or power electrodes E3 and E5 at the ends and two detection electrodes E1 and E2 just before the branch point V of the channel system. At the branching point V, a channel branches off, which in turn branches into three. At the ends there is a reservoir P, a mixing reactor R and another reservoir with a power electrode E4. If a mixture of substances along the

Separation channel T1 separated, can be determined using the detection electrodes E1 and E2, when the desired sample substance to the Branch point V of the channel system arrives. At this moment the potential is switched so that there is now a potential difference between E5 and E4. As a result, the selected substance migrates into the branch of the canal system. After the detection electrodes E1 and E2 indicate that the substance has passed the junction, the potential can be switched again. The substance secreted into the branch can now be transported mechanically by a liquid stream from the reservoir at E4 into the mixing reactor R. In addition, other substances, for example

Reactants for derivatization are passed into the mixing reactor, where they mix with the sample substance and, if necessary, react with it.

Figure 4 schematically shows an example of a miniaturized planar analysis system with an integrated device according to the invention for ejecting samples, a device for feeding defined sample volumes and a separation channel. Such an analysis system offers the possibility of giving up a defined large sample volume, for example separating it by means of ITP, by means of the invention

Discharge device to separate a certain fraction of the sample and optionally to separate and analyze the separated fraction or the rest of the sample again or to remove it from the system. The device for sample application consists of the channel sections K1 and K2, which are delimited by the fluid connections F1 and F2 or F2 and F3. Tightly closing micropumps or micropumps and valves are typically used as fluid connections. The volume of the channel section K1 is typically 5 or 10 μl, that of the channel section K2 0.5 or 1 μl. By opening the fluid connection F2 and simultaneously filling the channel section K1 with the sample solution via the fluid connection F1, the volume of K1 certain defined volume of the sample is filled into the channel system. A larger volume can be added if the fluid connection F3 is opened instead of the fluid connection F2. Then the given volume of the sample results from the sum of the volumes of the channel sections K1 and K2. If, on the other hand, the volume of the sample is to be smaller, only the channel section K2 is filled by opening the fluid connections F2 and F3. By varying the size of the channel sections K1 and K2 or also by adding further channel sections limited by fluid connections, the application volume can be varied and adapted to the corresponding requirements of the sample. The sample is separated in the subsequent channel system (K3, K4, K5). For this purpose, liquid or buffer reservoirs R1, R2 and R3 as well as power electrodes L1, L2 and L3 are located at the ends of the entire channel system, ie subsequently to K1, K4 and K5. The buffer reservoirs can be filled via the fluid connections F1, F4 or F5. If only channel section K1 is used for sample application, additional channel section K2 can be used to extend the separation distance. For purely analytical questions, the separation of the sample can be extended to channel section K3 up to channel section K5. The detection is then carried out by means of the detection electrodes D3 and D4 which are attached just before R3. If a fraction of the sample is to be separated from the rest, the device according to the invention is used for the removal. This is formed by the separating channel section K3, the branching point Vz, the two branches

Channel sections K4 and K5, the detection electrodes D1 and D2 located in front of the branch point Vz, and by a switching device (not shown in the figure) for controlling the power electrodes. As soon as the desired fraction passes through the detection electrodes D1 and D2 during the separation, the potential of the power electrodes L1, L2 and L3 can be modified accordingly. If the transport from L3 to L2 took place first, the fraction can be switched to a potential difference between L3 and L1 at the junction Vz can be discharged into the channel K4. After the fraction Vz has passed, the rest of the sample is transported back to K5 by switching again. The discharged fraction can then be removed from the analysis system via the fluidic connection F4. The rest of the sample remaining in K5 can be analyzed again via the detection electrodes D3 and D4. In the same way, the fraction to be discharged can be discharged by switching the power electrodes at Vz in channel section K5, while the rest of the sample is transported in channel section 4. In this case, the removed fraction at D3 / D4 can be detected again. There is also the option of filling the duct system with two different buffer systems and thus performing two different separations directly one after the other. For this purpose, the channel sections K3 and optionally additionally K2 are filled with the first buffer system. From the branch point Vz, the channel sections K4 and K5 are filled with the second buffer system. The first separation takes place along K2 / K3. At Vz, a fraction of the sample can then be discharged into channel section K5 or the entire sample can be transferred into this channel section. As soon as the sample reaches this channel section with the other buffer system, the second separation then takes place. The two separations are checked via the detection electrodes D1 and D2 for the first separation and the optional discharge, and with D1 / D2 for checking the second separation. In this way, for example, an isotachophoretic separation and an electrophoretic separation or two isotachophoretic separations can be combined.

Even without further explanations, it is assumed that a person skilled in the art can use the above description to the greatest extent. The preferred embodiments and examples are therefore only as descriptive disclosure, in no way to be construed as limiting in any way.

The complete disclosure of all of the applications, patents and publications mentioned above and below, and the corresponding application DE 199 27 535, filed on June 16, 1999, is incorporated by reference into this application.

Claims

Expectations

1. Device for discharging fractions for planar microstructured analysis systems, consisting essentially of a branched channel system, at least three transport electrodes, at least one detection device in front of a branching point of the channel system and an electrical switching device.

2. Device according to claim 1, characterized in that the detection device is an electrochemical detector.

3. Use of a device according to claim 1 or 2 in a planar, microstructured analysis system.