EP3740315A1

EP3740315A1 - Microfluidic device and method for the operation thereof

Info

Publication number: EP3740315A1
Application number: EP18845400.3A
Authority: EP
Inventors: Tino Frank
Original assignee: Robert Bosch GmbH
Current assignee: Robert Bosch GmbH
Priority date: 2018-01-15
Filing date: 2018-12-19
Publication date: 2020-11-25
Also published as: JP7171739B2; CN111565847A; DE102018200518B4; CA3088037A1; US20210114031A1; WO2019137775A1; DE102018200518A1; CN111565847B; KR20200110323A; JP2021510627A

Abstract

The invention relates to a method for operating a microfluidic device (1), comprising at least the following steps: a) providing at least one first medium (2, 9) at a first location of the microfluidic device (1), b) transporting at least one first medium (2, 9) from a first location to a second location of the microfluidic device (1), the at least one first medium (2, 9) being surrounded by at least one second medium (3) in such a way that the at least one first medium (2, 9) only borders on the at least one second medium (3) and on fluid boundaries (24) of the microfluidic device (1) or only on the at least one second medium (3), wherein the at least one first medium (2) and the at least one second medium (2, 9) cannot be mixed with one another.

Description

description

title

Microfluidic device and method of operation of the prior art

Microfluidic systems allow the analysis of small sample volumes with high sensitivity. Automation, miniaturization and parallelization of procedures allow a reduction of manual steps and can therefore help to avoid errors. Miniaturization of microfluidic systems also allows laboratory processes to be carried out directly at the sample, so that no general laboratory environment is needed.

Instead, a process can be reduced to a fluidic chip. Therefore, microfluidic applications may also be referred to as "lab-on-chip". This field of application of microfluidics is also referred to as "point-of-care (PoC)".

A particular challenge for microfluidic systems is the transfer of macroscopic samples into the microfluidic environment.

Disclosure of the invention

Here is a particularly advantageous method for operating a

Microfluidic device and a microfluidic device for this method presented. The dependent claims specify particularly advantageous developments of the method.

In particular, microfluidically limited sample solutions can be moved without loss in a microfluidic device with a microfluidic chamber and channel system using the described method.

In particular, with the described method a limited sample (For example, cell lysate from a few cells, cfDNA material, cytokine enrichment) from a sample input chamber into a chamber to perform a

Detection method (e.g., PCR) are transferred bubble and lossless.

Sample addition may allow rare material (e.g., cell-free DNA,

circulating cancer cells, secreted cytokines, lysate from a few cells) in a small volume of a microfluidic device are enriched and thus presented in a high concentration. These

Sample entry chamber must not be in the same location of the microfluidic device in the described method, in which a

Evaluation and / or further processing take place. The sample may instead be transported within the microfluidic device by the described method. Such transport often occurs in the prior art in an aqueous solution by laminar flow. However, this may result in material being deposited on the channel walls or thinned by more liquid or diffusion. Furthermore, through a contact surface to the outside world and / or by a lack of possibility for prewetting the chamber air into the system, which can lead to disturbing bubbles for subsequent processes.

The term "microfluidic" refers here primarily to the order of magnitude of the microfluidic device. The microfluidic device is characterized in that physical phenomena that are generally associated with microtechnology are relevant in the fluidic channels and chambers arranged therein. These include, for example, capillary effects, effects (especially mechanical effects) associated with

Surface tensions of the fluid are. Other effects include thermophoresis and electrophoresis. These phenomena are usually dominant in microfluidics over effects such as gravity. The

The microfluidic device can also be characterized in that it is made at least partially by a layer-by-layer process and that channels are arranged between layers of the layer structure. The term

"Microfluidic" may also be characterized via the cross-sections within the device which serve to guide the fluid. Typical are, for example, cross sections in the range of 100 pm [microns] times 100 pm up to 800 pm times 800 pm. Also much smaller cross sections, for example in the range of 1 mhh to 20 mhh [microns], in particular in the range of 3 mhh to 10 mhh are possible.

The microfluidic device may in particular be a so-called "lab on a chip" or a "point-of-care" system (PoC). Such a "lab on a chip" is designed and set up to perform biochemical processes. That means functionalities of a

macroscopic laboratories z. B. be integrated into a plastic substrate. The microfluidic device may e.g. As channels, reaction chambers, upstream reagents, valves, pumps and / or Aktuations-, detection and

Have control units. The microfluidic device can make it possible to process biochemical processes fully automatically. This can z. B. Tests on liquid samples are performed. Such tests can z. B. find application in medicine. The microfluidic device may also be referred to as a microfluidic cartridge. In particular, by introducing samples into the microfluidic device, biochemical processes can be carried out in the microfluidic device. The samples may also be admixed with additional substances which trigger, accelerate and / or facilitate biochemical reactions.

In particular, with the method described, a first medium can be transported from a first location of the microfluidic device to a second location of the microfluidic device.

In step a) of the described method, at least a first medium is provided at a first location of the microfluidic device.

The first medium is preferably a liquid, in particular an aqueous solution. In particular, the first medium may be a sample to be examined.

By "providing" is meant in particular that the at least one first medium is brought to the first location of the microfluidic device, for example by filling the at least one first medium through an opening in the microfluidic device. "Provision" also includes, for example, the microfluidic device already contained the at least one first medium before the beginning of the described method. So For example, a microfluidic device can be obtained from a supplier in which the at least one first medium is already arranged upstream in a chamber. It is also possible for the at least one first medium in step a) to be obtained by combining several substances and provided in this respect. For example, a solvent may be preceded in the microfluidic device. When a sample is added to the microfluidic device, the solvent may be added to the sample. The solution of the sample in the solvent may be the first medium.

In step b) of the described method, the at least one first medium from the first location to a second location of the microfluidic

Device transported. In this case, the at least one first medium of at least one second medium is enclosed in such a way that the at least one first medium only to the at least one second medium and to

Fluidbegrenzungen the microfluidic device or only adjacent to the at least one second medium. The at least one first medium and the at least one second medium are not miscible with each other.

In step b), the transport of the first medium takes place through the microfluidic device. In this case, the at least one first medium can be protected particularly well. In particular, the at least one first medium of the at least one second medium is preferably enclosed in such a way that the at least one first medium adjoins only the at least one second medium and optionally additionally fluid boundaries of the microfluidic device.

As fluid limitation comes here in particular every wall of the

Microfluidic device, for example, which limits a channel or a chamber of the microfluidic device. Media such as the at least one first medium and the at least one second medium can, within the microfluidic device in particular within the

Fluid boundaries are present and moving. In particular, the fluid boundaries may have a material such as glass and / or plastic on the fluid to be limited. The at least one first medium can in particular be prevented from coming into contact with other substances in step b). This can be achieved by the at least one first medium, insofar as it is not in contact with a fluid boundary, being in contact only with the at least one second medium. Characterized in that the at least one first medium and the at least one second medium are not miscible with each other, the at least one first medium can be transported without change by contact with the second medium. The at least one second medium can in particular be understood as an aid for the transport of the at least one first medium. After transport, the at least one first medium and the at least one second medium can be separated from one another.

The at least one second medium is preferably an oil. It is also preferred that the at least one second medium is an organic substance.

In particular, it is preferred that the at least one first medium is polar and the at least one second medium is nonpolar. This is the case, for example, with water as the first medium and oil as the second medium. As an aqueous solution, water can be added with classical attributes such as Tween, Triton-X, BSA and / or calcium for the first medium. In particular inert mineral oils, silicone oils and / or fluorinated oils can be used as possible second media. On the use of surfactants is

preferably omitted.

In particular, with the described method, a defined volume of an aqueous phase (as the at least one first medium) in an oil phase (as the at least one second medium) can be trapped and moved in a controlled manner. For example, an analyte present in the aqueous phase can be present in a limited, small amount and can be processed without loss and dilution in the microfluidic device.

By using the at least one second medium (in particular an organic phase), the at least one first medium (in particular an aqueous volume) can be enclosed in such a way that, for example, a limited analyte in the at least one first medium does not dilute by deposition or diffusion. It is thus in particular a loss-free transport of Limited sample materials (as the first medium) possible. For example, a lysate produced locally in a microfluidic small volume may be transported from a few cells from an input chamber to another location in the microfluidic device for biochemical processing.

Lossless transport of limited material such as DNA, proteins, and / or single cells may allow for a design of a microfluidic processing unit, for example, where a heater or optical units are provided at a location other than a trial input. This may allow a particularly universal design of the microfluidic device.

Furthermore, by a first filling of the microfluidic device with the at least one second medium, a wetting of fluid boundaries (ie in particular of channel walls and / or chamber walls) can take place. In this case, a thin layer of the second medium can deposit on the fluid boundaries. In the case of polycarbonate, for example, this thin layer may have the advantage that no DNA is bound to the polycarbonate (or to the layer of the second medium) as material of the fluid boundary. This may contribute to a lossless transport of DNA in the at least one first medium.

The microfluidic device preferably comprises a one-way flow system which enables a diagnosis in the manner of point-of-care. The components of the microfluidic device can thereby in a

Polycarbonate injection molded part are manufactured.

In a preferred embodiment of the method, in step a), a predeterminable volume of the at least one first medium is provided in a chamber of the microfluidic device, wherein the chamber has at least one connection, and wherein the predeterminable volume of the at least one first medium in the chamber is separated and is measured by the at least one port is flowed around with the at least one second medium outside the chamber.

The at least one first medium may in particular be a sample to be analyzed. In particular, in such a case, it may be advantageous to have a precisely determined amount (in particular a precisely defined volume) of the for example, to use at least a first medium for analysis. In order to obtain such an accurately determined amount of the at least one first medium, the desired amount of the at least one first medium according to the present embodiment can be separated and measured.

Thus, in particular, the chamber of the microfluidic device considered in this embodiment can be filled, in particular via the at least one connection, with the at least one first medium. If the chamber is completely filled with the at least one first medium, the volume of the at least one first medium corresponds to the (preferably known) volume of the chamber. However, a demarcation between the volume within the chamber and outside the chamber, especially in the region of the at least one port, can be problematic. It may therefore be unclear where exactly the boundary of the chamber passes through the at least one connection.

In the present embodiment, this limit can be determined by the at least one second medium. Preferably, the at least one connection is designed in such a way that a flow of the at least one second medium (with preferably fixed parameters, such as, for example, a flow velocity) flows around the at least one connection in a reproducible manner. In the case of such a reproducible recirculation, there results an interface between the at least one first medium and the at least one second medium, which is located in particular in the region of the at least one connection. By this limit, the volume of the chamber can be clearly defined.

In a further preferred embodiment of the method, a plurality of first media are provided in step a), wherein the plurality of first media according to step b) are transported such that the plurality of first media are mixed in a chamber of the microfluidic device.

In particular, the plurality of first media may be components of a substance to be analyzed. In this case, the plurality of first media may remain separated from one another, for example, until an analysis is to be carried out. For example, a reaction between the plurality of first mediums may be prevented until the analysis is performed.

The two-phase technology (in which the at least one first medium and the at least one second medium are used) can be used in particular to mix two limited sample volumes or fluids (as the first multiple media). Both preferably aqueous volumes of the first media can be guided lossless to a chamber and there

be mixed by diffusion. Such mixing can therefore be carried out sufficiently fast, in particular, if the volumes to be mixed are sufficiently small.

In a further preferred embodiment of the method, at least a portion of the at least one first medium and / or at least a portion of the at least one second medium in step b) are transported at least temporarily by peristaltic pumping.

By peristaltic pumping is meant here a pump which promotes a fluid by means of peristalsis. A typical peristaltic pump is a peristaltic pump, also called peristaltic pump. Peristaltic pumps are displacement pumps in which the medium to be pumped is pushed through a channel by an external mechanical deformation.

Microfluidic peristaltic pumps may be constructed by a plurality of valves. Frequently used microfluidic valves include a channel which is closable by a movement of the channel wall due to an electrical force or a magnetic force. Such valves produce an (internal) volume change of the channel. If such valves in one

Series connection are arranged along a channel can be achieved by a suitable control of the valves that a peristaltic of the channel occurs, which causes a promotion of the liquid. By opening and closing the valves occur volume changes of a channel through which a transport of a medium through the channel of the microfluidic device takes place. A peristaltic pump has the advantage that it requires no (other) pumping elements in addition to the valves (for example mechanically or electrically operating pump chambers). It is sufficient that the majority of the valves is provided. For peristaltic pumping it is preferred that there is the possibility of an (automatic) valve circuit, according to which the valves are automatically controlled in a suitable order for the promotion.

By combining fixed channel geometries with on-chip pumps, dynamic volume adjustment can be achieved. That is in particular in a two-phase system (comprising the at least one first medium and the at least one second medium) particularly well possible. By contrast, if, for example, only one aqueous phase were provided (that is, for example, only the at least one first medium), then the volumes would be predetermined by fixed geometries of the chambers. With a two-phase system, however, the chamber geometry only forms an upper limit of the possible volume. Since an aqueous and an oil phase do not mix, the inert oil can compensate for the volume not needed by the aqueous phase. This may allow additional dynamic component and volume adjustment within the microfluidic device.

In a further preferred embodiment, the method further comprises at least the following method step, which is carried out before, during or after step b):

c) removing at least one gas inclusion.

Gas inclusions can occur especially in the form of gas bubbles in the

microfluidic device. Gas inclusions may be particularly disadvantageous because of these volumes can be determined only inaccurate and / or because it can cause reactions between the gas and in particular the at least one first medium. In the present embodiment, gas inclusions can be removed. The gas may be, in particular, air. Also, the gas can be a product of chemical reactions.

The at least one gas inclusion can be removed in particular by transporting a medium. In particular, a medium enclosing the at least one gas inclusion can thus be replaced by the microfluidic one

Be moved device that the at least one gas enclosure reaches a location of the microfluidic device, on which a (opposite

Gravitational direction) upwardly directed flow path for the escape of the gas for the at least one gas inclusion is accessible.

The at least one gas inclusion can be removed, in particular, insofar as the gas is passed out of the microfluidic device or that the gas is introduced into at least one part of the microfluidic device other part of the microfluidic device is passed, wherein the gas is less harmful or disturbing in the latter part.

However, the at least one gas inclusion can also be removed by removing a gas dissolved in the at least one first medium (that is to say a substance which is present in gaseous form under normal conditions) from the at least one first medium. Thus, a two-phase system (comprising the at least one first medium and the at least one second medium) may particularly facilitate degassing of the microfluidic device because many oils (which are preferably used as the at least one second medium) have a higher gas solubility than water (the preferably as an essential component, the at least one first medium is used). Therefore, it can be achieved that dissolved in water gases in the

Go over the gas phase and then dissolved in the oil again.

In that regard, gas containment can also be removed by quasi-phase extraction.

Removal of the at least one gas inclusion may be achieved, in particular, in the preferred embodiment of the method in which the microfluidic device is oriented, at least during a portion of step c), such that one side of a portion from which the at least one gas inclusion is removed a horizontal plane is tilted.

Preferably, the microfluidic device is oriented during the entire step c) in such a way that one side of a section from which the at least one gas inclusion is removed is tilted relative to a horizontal plane. Preferably, the microfluidic device is oriented such that the side of the portion from which the at least one gas inclusion is removed is tilted with respect to the horizontal plane by an angle in the range of 20 ° to 45 °, in particular by 30 °.

By tilting the microfluidic device, it can be achieved that the gas can escape from the at least one gas inclusion upwards (ie counter to the gravitational direction). In a further preferred embodiment of the method, a temperature of a fluid in which the at least one gas inclusion is enclosed is changed in step c).

By the temperature change, a solubility of the gas in the fluid can be reduced, so that the gas from the gas inclusion can escape more easily.

In a further preferred embodiment of the method, the at least one gas inclusion in step c) is removed by transport of the at least one first medium and / or the at least one second medium.

The use of the at least one first medium and / or the

At least one second medium for removing the at least one gas inclusion may be particularly advantageous in that the at least one first medium and / or the at least one second medium are present anyway in the microfluidic device or that these medium are moved anyway by the described method ,

In a further preferred embodiment of the method, a shuttle polymerase chain reaction [shuttle PCR] is carried out, wherein the at least one first medium is a reaction medium of the shuttle polymerase chain reaction.

PCR is a method of amplifying DNA using the enzyme DNA polymerase. A PCR can in particular under

Change in the temperature of the reaction medium take place. In a shuttle PCR, this temperature is changed by moving the reaction medium between locations with different temperatures (in particular between

Chambers with different temperature) is transported. So a temperature change can be done very quickly.

The described method may in particular provide the advantage that a shuttle PCR can be defined, bubble-free and carried out without loss. In a single-phase system, on the other hand, there would be the risk that residues of the reaction medium could remain in a channel and / or air could enter the reaction chamber. As a further aspect, a microfluidic device is provided, which is intended and arranged for carrying out the method described.

The particular advantages described above and

Design features of the method are described on

Microfluidic device applicable and transferable.

Further details of the invention and embodiments, to which the invention is not limited, will be explained in more detail with reference to the drawings. Show it:

Fig. La to ld: four schematic representations of microfluidic

Devices with a first medium and a second medium

2a to 2c: schematic representations of a microfluidic

Device in three successive times, wherein a volume of a first medium separated and

is measured,

3a to 3e: schematic representations of a microfluidic

Device in five successive times, wherein a first medium is generated and a volume of the first medium is separated, measured and transported away,

4a to 4e: schematic representations of a microfluidic

Device in five successive times, with a first medium separated into two sub-volumes, measured and transported away,

Fig. 5: a schematic representation of a microfluidic

Device in which a chamber is partially filled with a first medium, 6a to 6f: schematic representations of a microfluidic

Device in six consecutive times, with two first media mixed together,

Fig. 7a to 7f: schematic representations of a microfluidic

Multi-valve device, differently connected for peristaltic pumping in six consecutive times,

8a and 8b: two schematic representations of the peristaltic

Pump,

9a to 9d: schematic representations of a microfluidic

Device at four consecutive times, performing peristaltic pumping,

10a to 10d: schematic representations of a microfluidic

Device in four consecutive times, with gas entrapment removed,

11: schematic representations of a microfluidic

Device that is tilted oriented,

12a to 12c: schematic representations of a microfluidic

Device in three consecutive times, with gas entrapment removed,

13a to 13d are schematic representations of a microfluidic

Device in four consecutive times, wherein a shuttle PCR is carried out, and

14 shows a schematic illustration of a method for operating a microfluidic device according to one of the exemplary embodiments from the previous figures.

In Fig. La to ld is shown as in a microfluidic device 1, a liquid phase as the first medium 2 between two oil phases as the second Medium 3 can be included. In this case, the first medium 2 can be presented in particular with a defined volume. The volume of the first medium 2 can be determined by the fixed and precisely producible geometry of the microfluidic device 1. In this form, therefore, a defined concentration of the first medium 2, in particular if I it is an analyte, are submitted.

In order to move the volume of the first medium 2 without the analyte diluting therein by diffusion, the first medium 2 is trapped between the second medium 3 (shown here by two oil phases). Since oil and water do not mix, there is no dilution of the first medium 2 by diffusion. This can enable a loss-free microfluidic transport of a defined volume of the first medium 2 through a first channel 5 or out of a first chamber 4. The first chamber 4 is connected via a first connection 25 to a first channel 5 and via a second connection 26 to a second channel 6. The first medium 2 and the second medium 3 are surrounded by fluid boundaries 24.

In particular, the first medium 2 may be arranged upstream in the first chamber 4 (FIG. 1 a) and transported through the first connection 25 from the latter into the first channel 5 (FIG. 1 b). In the further course of the first channel 5, the first medium 2 can be enclosed between the second medium 3 (FIG. 1c).

In Fig. Id, another situation is shown, in which the first medium 2 is enclosed in a first chamber 4 of the microfluidic device 1.

FIGS. 2 a to 2 c and 3 a to 3 e show two embodiments of a microfluidic device, with which a defined volume of an aqueous phase as the first medium 2 can be brought between two inert oil phases as the second medium 3.

In the embodiment according to FIGS. 2 a to 2 c, a first chamber 4 of known volume is arranged between a first channel 5 and a second channel 6 parallel to the first channel 5. The first chamber 4 is connected via a first connection 25 to a first channel 5 and via a second connection 26 to a second channel 6. By suitable

Fluid control (for example, by using valves or a

Pressure compensation system), the flow can be in different directions and Channel compositions are set (for example, as a flow only in the channels 5, 6 or as a flow from the second channel 6 through the first chamber 4 in the first channel 5). In a first step, the first chamber 4 (FIG. 2 a) initially filled with a second medium 3 is flowed through with an aqueous phase as the first medium 2 and the flow is stopped so that the first

Chamber 4 is completely filled with the first medium 2 (Fig. 2b). The adjacent channels 5, 6, but not the first chamber 4, are then flushed with oil as the second medium 3, so that the first chamber 4 is surrounded by two oil-filled channels 5, 6 (FIG. 2c). Now, a flow from the second channel 6 through the first chamber 4 in the first channel 5 can be adjusted. In this case, the three phases (ie the second medium 3 in the second channel 6, the first medium 2 in the first chamber 4 and the second medium 3 in the first channel 5) move laminarly without mixing.

In the second embodiment according to FIGS. 3 a to 3 e, the first chamber 4 only adjoins (via a first connection 25) a first channel 5 and not a first channel 5 and a second channel 6 as in the first embodiment

Embodiment according to FIGS. 2a to 2c. A portion of the first chamber 4 is open or (as shown) through a gas-permeable membrane 7 of the

Separated environment of the microfluidic device 1, so that exchanged between the first chamber 4 and the environment air and / or pressure can be compensated. The gas-permeable membrane 7 can

in particular be used as a sample entry area, in particular for applications in which only small amounts of a sample are introduced into the microfluidic device.

In Fig. 3a is also shown that in the first chamber 4, a sample. 8

(For example, as a solid or Lyobead) is present. To release the sample 8 and / or to fill the first chamber 4, the first channel 5 is first filled with oil as the second medium 3 in order to vent the entire system. The first chamber 4 is then filled with a liquid phase as the first medium 2 (FIG. 3b). When the chamber is completely filled, the first channel 5 is again flowed through with oil as the second medium 3 and thus the first chamber 4 is completely closed (FIG. 3c). The first medium 2 can then again be enclosed between an oil phase as a second medium 3, by the first

Chamber 4 is pumped out via the first channel 5 (Figure 3d) until the first chamber 4 is empty (Figure 3e). A further exemplary embodiment of a microfluidic device 1 with which a defined volume of an aqueous phase as the first medium 2 can be brought between two inert oil phases as the second medium 3 is shown in FIGS. 4 a to 4 e. In contrast to the embodiment of FIGS. 2a to 2c, two partial volumes of the first medium 2 are removed from the first chamber 4 in succession. The starting point shown in Fig. 4a corresponds largely to the representation in Fig. 2c. By a flow of the second medium 3 from the second channel 6 into the first channel 5, part of the first medium 2 is removed from the first chamber 4 (FIG. 4b). Subsequently, the first channel 5 is again flowed through by the second medium 3 (FIG. 4c). Thereafter, the remaining part of the first medium 2 is removed from the first chamber 4 (Figure 4d). As can be seen in FIG. 4e, in the further course of the first channel 5, the first medium 2 is present in two parts, which are each enclosed by the second medium 3. FIGS. 4a to 4e thus show that a two-phase system (with first medium 2 and second medium 3) can also be utilized in order to fill a microfluidic chamber only partially bubble-free with an aqueous phase (as first medium 2). The residual volume of the chamber can be compensated accordingly with an inert oil phase (as the second medium 3). This can be a dynamic adaptation of

Allow reaction volume.

FIG. 5 shows a state from the exemplary embodiment from FIGS. 4a to 4e, in which the first chamber 4 is partially filled with the first medium 2 and partly with the second medium 3.

FIGS. 6a to 6f show an exemplary embodiment of a microfluidic device

Device 1, in which two aqueous phase fluids (as a first medium 2 and a further first medium 9) are mixed. In this case, the first medium 2 is half filled in a first chamber 4 with a defined volume (Fig. 6a). The first chamber 4 is connected via a first connection 25 to a first channel 5 and via a second connection 26 to a second channel 6. The feeding first channel 5 is then completely filled with oil as the second medium 3 again (FIGS. 6b and 6c). Then the first channel 5 is filled with the further first medium 9 (FIG. 6d). With the further first medium 9 then the first chamber 4 is filled (preferably slowly), so that the whole of the first chamber 4 with the two first media 2, 9 in the correct ratio is filled. The first channel 5 is then filled again with the second medium 3, so that the two first media 2, 9 are again enclosed in the first chamber 4 by the second medium 3 in the channels 5, 6 (FIG. 6e). By

Diffusion can mix the two first media 2, 9 in the first chamber 4 quickly, especially if the respective volumes are small. The result is shown in Fig. 6f, in which in the first chamber 4, a mixture 10 of the two first media 2, 9 is present. The mixing process can be accelerated by a temperature change. If desired, (bio) chemical reactions can also be carried out during mixing. The

Combination of defined pumping processes and predetermined

Chamber geometries may allow for mixing with different ratios between the first media 2, 9.

FIGS. 7a to 7f show how fluids in a linear or circular channel system of a microfluidic device can be moved by means of valves at a controlled speed. Valves in the microfluidic

Devices can not only be used to open and close microfluidic paths, but can also be used as peristaltic pumps. Along a desired microfluidic pathway (which may be linear or circular and shown here as a linear first channel 5), the valves form a peristaltic pump by serially opening and closing. 7a to 7f show the principle using the example of three adjacent valves 11, 12, 13. A circle indicates an open valve, while a cross indicates a closed valve. The valve status can also be displayed digitally, for example by setting a "1" to "open" and a "0" to "closed". The valve status sequence 100 (Figs. 7e), 110 (Fig.

7d), 010 (Fig. 7c), 011 (Fig. 7b), 001 (Fig. 7a), 101 (Fig. 7f) generates a movement from left to right in the illustration shown. The sequence 001 (Fig. 7a), 011 (Fig. 7b), 010 (Fig. 7c), 110 (Fig. 7d), 100 (Fig. 7e), 101 (Fig. 7f) produces a quasi-laminar flow from the right to Left.

It is also shown in FIGS. 8a and 8b that the valves 11, 12, 13 do not have to be placed next to one another, but can be arranged arbitrarily along the first channel 5. This has the advantage that no special pump valves need to be placed, but already existing valves 11, 12, 13 can be used. The flow speed can be in one determined interval by means of a duration of a pause between successive valve positions.

FIGS. 9a to 9d show how the embodiment according to FIGS. 2a to 2c can be realized by a peristaltic pump in a multi-chamber system (comprising a first chamber 4, a second chamber 14 and a third chamber 15). In a first step, the microfluidic device 1 is completely filled with an oil phase as the second medium 3 (FIG. 9a). This does not necessarily have to be done by peristaltic pumping. When the microfluidic device 1 is filled, a fifth valve 18 and a sixth valve 19 are closed between the chambers 4, 14, 15 and a first channel 5 (FIG. 9b). A laterally to the chambers 4, 14, 15 arranged channel 5 is then at least partially filled with the first medium 2. If this state is reached, the fifth valve 18 and the six valve 19 are opened to the chambers 4, 14, 15 and it is so long with the valves 11, 12, 13 pumped peristaltically until the first chamber 4 completely with the first medium 2 is filled (Figures 9c and 9d). The pump can be coupled to an optical feedback system and automatically stopped when fully charged. Alternatively, even a trapped aqueous plug with the chamber volume can be introduced into the first chamber 4. If the first chamber 4 is completely filled, the fifth valve 18 is closed to the first chamber 4 and the first channel 5 again completely flushed with the second medium 3 (by opening a fourth valve 17), so that only the first medium 2 in the first chamber 4 remains (Figure 9d).

FIGS. 10 a to 10 d show how a two-phase system (with a first medium 2 and a second medium 3) can be used to remove gas inclusions 16 in the first medium 2. Gas inclusions 16, such as perturbing bubbles, are removed by a temperature gradient in this design. For this purpose, three microfluidic chambers 4, 14, 15 are arranged one behind the other and connected to each other by means of a respective small channel. Each of the three chambers 4, 14, 15 is individually heated. The first chamber 4 is set to the highest temperature and the second chamber 14 and the third chamber 15 to a deeper. In this case, the heated gas inclusions 16 move from the first medium 2 into the colder second medium 3. If the gravitational force continues to act along the chamber geometry (as shown), the

Gas inclusions 16 due to the lower density all the way up. It forms Thus, a fluid system in which the first medium 2 in the first

Chamber 4 is present, wherein the second chamber 14 and the third chamber 15 are filled with the second medium 3. In the third chamber 15, a gas phase can form. The temperature can then be raised again in the second medium 3 to ensure that the gas settles at the top. The phase system can be displaced in each case by one chamber, the first medium 2 then being located, for example, in the second chamber 14 and the bubble-free second medium 3 in the third chamber 15. The blister-containing first chamber 4 is then eliminated from the chamber system. The first chamber 4 can be closed or refilled with the second medium 3.

11 shows how an inclination of the microfluidic device 1 and thermally different zones for the removal of gas inclusions 16 according to FIGS. 10a to 10d can be utilized. The gravity does not have to work completely. Instead, an inclination (eg of 30 °) is possible. FIG. 11 shows a horizontal plane 21 and an angle 22 between the horizontal plane 21 and a side 23 of the microfluidic device 1 from FIGS. 10a to 10d. The three-chamber system shown in FIGS. 10a to 10d may be oriented such that the first chamber 4 is located below the first medium 2 (ci in FIG. 11). Each chamber 4, 14, 15 is then in its own thermal zone (Ti, T ₂ , T ₃ ). Gravity may cause the light gas to move from the gas enclosure 16 to the upper end of the third chamber 15 by heating (C3 in FIG. 11).

FIGS. 12a to 12c show an embodiment in which a gas enclosure 16 as in FIGS. 10a to 10d and 11 can be removed. In FIGS. 12a to 12c, it is assumed that the gas inclusion 16 according to the

Method according to FIGS. 10a to 10d has been removed from the first medium 2 with the aid of the conditions from FIG. 11 (FIG. 12a). In order to remove the gas inclusion 16 from the microfluidic device 1, a second medium 3 is pushed from below through the three chambers 4, 14, 15 until the first medium 2 has been completely displaced from the first chamber 4 into the second chamber 14 , In this case, the gas inclusion 16 is displaced from the third chamber 15 into the first channel 5 (FIG. 12b). Subsequently, through

Moving the second medium 3 through the first channel 5 (but not through the chambers 4, 14, 15), the gas from the microfluidic device first be discharged, leaving a completely bubble-free microfluidic

Device 1 remains (Fig. 12c).

Figs. 13a to 13d show how with a two-phase system and the

Combination of the above-described removal of a gas inclusion 16 bubble-free shuttle PCR can be performed. In a shuttle PCR, unlike a thermal cycler, the temperature of heaters is not dynamically altered but the reaction mixture is shifted between different heaters at constant temperatures. In order to carry out a shuttle PCR, the microfluidic device 1 can be designed in particular according to FIGS. 7a to 7f, 10a to 10d and 11. In the arrangement of three chambers 4, 14, 15 and a first channel 5, three valves 11, 12, 13 are preferably provided which form a peristaltic pump. The PCR reaction mixture (as the first medium 2) is preferably introduced bubble-free in the first chamber 4, while the second chamber 14 and the third chamber 15 and the first channel 5 are filled with the second medium 3 (Fig. 13a). The chambers 4, 14, 15 are set to the appropriate temperatures required for the PCR. The first medium 2 as a PCR mixture can then be used for respective times in the corresponding chambers 4,

14, 15 before it is peristaltically pumped into the next of the chambers 4, 14, 15 (Figures 13b to 13d). In particular, it shows how to shuttle back and forth between two temperatures (commuted, engl., "Shuttled"). By reversing the pumping frequency, as described with reference to FIGS. 7a to 7f, it is possible to pump in both directions.

14 shows a method for operating a microfluidic device 1 according to one of the exemplary embodiments from the previous figures. The method comprises the following steps:

a) providing at least one first medium 2, 9 at a first location of the microfluidic device 1, b) transporting the at least one first medium 2, 9 from the first location to a second location of the microfluidic device 1, wherein the at least one first medium 2 9 is surrounded by at least one second medium 3 such that the at least one first medium 2, 9 only to the at least one second medium 3 and fluid boundaries 24 of the microfluidic device 1 or only to the at least one second Medium 3 adjacent, and wherein the at least one first medium 2, 9 and the at least one second medium 2, 9 are not miscible with each other.

Furthermore, the method preferably comprises the following (dashed line) method step, which is carried out before, during or after step b):

c) removing at least one gas inclusion 16.

In the example of FIG. 14, step c) is performed after step b).

Claims

claims

A method of operating a microfluidic device (1) comprising at least the following steps:

a) providing at least one first medium (2, 9) at a first location of the microfluidic device (1),

b) transporting the at least one first medium (2, 9) from the first location to a second location of the microfluidic device (1), wherein the at least one first medium (2, 9) is surrounded by at least one second medium (3) in that the at least one first medium (2, 9) adjoins only the at least one second medium (3) and fluid boundaries (24) of the microfluidic device (1) or only the at least one second medium (3), and wherein the at least one first medium (2, 9) and the at least one second medium (2, 9) are not miscible with each other.

2. The method of claim 1, wherein in step a) a predeterminable volume of the at least one first medium (2, 9) in a chamber (4, 14, 15) of the microfluidic device (1) is provided, wherein the chamber (4, 14, 15) has at least one connection (25, 26), and wherein the

predetermined volume of the at least one first medium (2, 9) in the chamber (4, 14, 15) is separated and measured by the at least one port (25, 26) with the at least one second medium (3) outside the chamber ( 4, 14, 15) is flowed around.

3. The method according to any one of the preceding claims, wherein a plurality of first media (2, 9) are provided in step a), and wherein the plurality of first media (2, 9) according to step b) are transported such that the plurality of first media ( 2, 9) in a chamber (4, 14, 15) of

Microfluidic device (1) are mixed.

4. The method according to any one of the preceding claims, wherein at least a portion of the at least one first medium (2, 9) and / or at least a portion of the at least one second medium (3) in step b) are transported at least temporarily by peristaltic pumping.

5. The method according to any one of the preceding claims, further comprising at least the following method step, which is carried out before, during or after step b):

c) removing at least one gas inclusion (16).

6. The method according to claim 5, wherein the microfluidic device (1)

at least during part of step c) is oriented such that one side (23) of a portion from which the at least one gas occlusion (16) is removed is tilted with respect to a horizontal plane (21).

7. The method according to any one of claims 5 or 6, wherein in step c) a

Temperature of a fluid in which the at least one gas inclusion is included, is changed.

8. The method according to any one of claims 5 to 7, wherein the at least one gas inclusion (16) in step c) by transport of the at least one first medium (2, 9) and / or the at least one second medium (3) is removed. 9. Method according to one of the preceding claims, wherein a shuttle

Polymerase chain reaction [PCR] is carried out, wherein the at least one first medium (2,

9) is a reaction medium of the shuttle polymerase chain reaction.

10. A microfluidic device (1) which is intended and arranged for carrying out a method according to one of the preceding claims.