Microfluidic device, biomarker detection device with microfluidic device and method

Abstract

A microfluidic device (12) for applying a sample of body fluid to a conditioned sensor element for the detection of molecular biomarkers in the body fluid and / or for conditioning the sensor element (14) using a conditioning fluid comprises at least one interaction chamber (40) that is partially closed by a sensor element (14). A pneumatic unit (70) is connected to the at least one first receptacle (46) and is configured to pressurize it and pump the body fluid or the conditioning fluid once and unidirectionally through the at least one interaction chamber (40) along the treated surface (16) and subsequently into the at least one disposal container (52), so that, in the case of body fluid, biomarkers adhere to the surface (16) or, in the case of conditioning fluid, the surface is conditioned.Furthermore, a biomarker detection device (18) and a method for applying a sample of body fluid to a conditioned sensor element (14) for biomarker detection and / or for conditioning a sensor element (14) for biomarker detection are proposed.

Description

[0001] The invention relates to a microfluidic device for applying a sample of body fluid from a human or animal body, but in particular from a human body, to a sensor element for biomarker detection and / or for conditioning a sensor element for biomarker detection. The invention further relates to a biomarker detection device comprising the microfluidic device and a method for applying the sample of body fluid to a conditioned sensor element or for conditioning a sensor element for biomarker detection.

[0002] The present invention is intended to improve, in general, the possibilities of detecting so-called diagnostic, prognostic and / or predictive biomarkers in body fluids (including, among others, intestinal fluid, ocular fluid, lung fluid, blood or cerebrospinal fluid), i.e., analyzing their presence in order to draw conclusions about diseases in humans or animals, including for the diagnosis of Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, diabetes, Huntington's disease, prion disease or a tumor in a patient.

[0003] Alzheimer's disease, also known as Alzheimer's, is a currently incurable neurodegenerative disorder in which nerve cells in the brain progressively die, leading to personality and behavioral changes in affected individuals. Millions of people worldwide are affected by Alzheimer's.

[0004] Early diagnosis of Alzheimer's disease is crucial for its progression, as while it is not yet curable, its steadily worsening course can be slowed by early intervention. The earlier treatment begins, the more beneficial it is. Therefore, early and reliable diagnosis presents a significant challenge in diagnostic medical technology, enabling timely intervention and slowing of the disease's progression. Prediagnostic methods play a particularly important role, as they allow for the prediction of the risk of a clinical Alzheimer's diagnosis and enable therapeutic interventions to begin even before the clinical onset of symptoms.

[0005] Established prediagnostic methods for predicting the risk of Alzheimer's disease rely on the analysis of specific protein defects that develop in the human body decades before the clinical onset of Alzheimer's. Analyzing these protein defects allows for predictions about the likelihood of developing Alzheimer's. From a scientific perspective, it is assumed that in the early stages of Alzheimer's, certain proteins, including β-amyloid (Aβ) peptides, misfold. According to current research, this misfolding and the resulting aggregation of peptides into beta-sheet-rich amyloid plaques is crucial for the course and progression of Alzheimer's.

[0006] For this reason, the presence of misfolded β-amyloid (Aβ) peptides in the blood plasma of test subjects, for example, is a sign of possible Alzheimer's disease and can be an important indicator for predicting the risk of developing Alzheimer's. Thus, misfolded β-amyloid (Aβ) peptide represents a candidate biomarker peptide that can be used as a biomarker in prediagnostic methods.

[0007] In the field of protein diagnostics, a number of methods exist for detecting candidate biomarker peptides in samples. For example, methods such as enzyme-linked immunosorbent assays (ELISA), surface plasmon resonance spectroscopy (SPR), or surface-based fluorescence intensity distribution analysis (sFIDA) can be used. However, it has been shown that these methods only provide indirect information about structural defects or misfolding of proteins. Furthermore, these methods are very complex in terms of sample preparation and execution and are designed for low sample throughput, making them unsuitable for large-scale clinical applications.

[0008] Another method in the field of protein diagnostics is the immuno-infrared sensor assay (hereinafter referred to as immuno-assay), which allows for the direct measurement of misfolded β-amyloid (Aβ) peptides in body fluids such as blood, blood plasma, blood serum or cerebrospinal fluid without prior isolation and preparation of candidate biomarker peptides.

[0009] An example of such an immunoassay is disclosed in US 2020 / 0141866A1. The described immunoassay focuses in particular on the protein diagnostics of misfolded amyloid-β(Aβ) peptides for the risk prognosis of Alzheimer's disease. The basic principle of the method is to bind an antibody to a sensor surface-modified with silanes or thiol linkers, which in turn can specifically bind candidate biomarker peptides based on (Aβ) structures.

[0010] In a first preconditioning step, a sensor crystal, for example, an ATR (attenuated total reflection) sensor crystal such as a germanium or silicon single crystal, is surface-treated with conditioning fluids containing silane or thiol linkers. In a second antibody conditioning step, the sensor surface is treated with an antibody-containing conditioning fluid. In a third sample deposition step, the sensor is then surface-treated with a sample fluid from a test subject, such as blood plasma, blood, or blood serum, whereby candidate biomarker peptides based on (Aβ) structures bind to the antibody-coated sensor surface.In a fourth step, the measurement step, the sensor is irradiated with an IR laser that passes through it via multiple internal total reaction, thereby recording an IR spectrum of the candidate biomarker peptides bound to the sensor surface and based on (Aβ) structures. The IR spectrum recorded by the bound candidate biomarkers is then evaluated, for example, by quantifying the shift of the IR bands of the (Aβ)-based candidate biomarker peptides with comparative measurements, in order to make a risk prediction regarding the potential onset of Alzheimer's disease in relation to the sample under investigation.

[0011] However, such immunoassays for Alzheimer's risk prediction are in an early stage of development and have so far only been conducted on a laboratory scale and as conceptual trials. In particular, developing such an immunoassay from the laboratory scale to industrial, large-scale medical application presents an immense challenge.

[0012] Known immunoassays use individual devices to perform the above-mentioned steps, which have a number of disadvantages that will be discussed below.

[0013] Typically, known immunoassays are based on a flow cell with a fluid port at each of its opposite ends. Both ports are fluid-flow connected to a common reservoir, so that the sample to be analyzed is pumped along a circuit between the flow cell and the reservoir via a peristaltic pump.

[0014] However, flow cells are expensive and require extensive preparation. Therefore, time-efficient throughput of larger sample volumes is not possible, and analysis is limited to individual samples.

[0015] The use of a peristaltic pump also increases the risk of contaminating the sample fluid. To avoid this, it is necessary to clean the peristaltic pump regularly, for example by autoclaving it, to prevent cross-contamination between different sample fluids.

[0016] Furthermore, the manual setup and dismantling of the experimental setup between measurements makes the reproducibility of the immunoassay more difficult and thus hinders efficient sample throughput.

[0017] Furthermore, the conditioning of the sensor surface and the subsequent sample application with the sample fluid do not take place in the same flow cell, meaning that several specialized devices are used for sample preparation and measurement. This procedure is time-consuming and increases the risk of sample fluid contamination, as the sensor is passed back and forth between the devices for conditioning and sample application.

[0018] Parkinson's disease and amyotrophic lateral sclerosis (ALS) should also be able to be predicted early by analyzing certain protein defects in the blood. The invention should therefore include, among other things, a device and a method for conditioning sensor elements and analyzing protein defects using these sensor elements for these diseases; that is, more generally, for preparing and carrying out an analysis of protein defects.

[0019] Therefore, the object of the present invention is to eliminate at least one disadvantage known from the prior art.

[0020] The invention provides a microfluidic device for applying a sample of body fluid to a conditioned sensor element for the detection of molecular biomarkers in the body fluid and / or for conditioning the sensor element by means of a conditioning fluid, comprising at least one interaction chamber, which is partially closed by a sensor element, wherein a first fluid channel opens at one end of the at least one interaction chamber and a second fluid channel opens at the opposite end, at least one first inlet for receiving the body fluid or the conditioning fluid, which is fluidically connected to the first fluid channel at one end of the at least one interaction chamber forming a fluid inlet, wherein the second fluid channel leads from its associated interaction chamber to a disposal container, wherein the sensor element has a sensor surface facing the interaction chamber, to which biomarkers adhere in the conditioned state, and comprising a pneumatic unit connected to the at least one first recording device and configured to pressurize it and pump the body fluid or conditioning fluid once and unidirectionally through the at least one interaction chamber along the treated surface and subsequently into the at least one disposal container, so that, in the case of body fluid, biomarkers adhere to the surface or, in the case of conditioning fluid, the sensor surface is conditioned.

[0021] The device according to the invention provides that when the already conditioned sensor element comes into contact with the body fluid sample, the sample is pumped through the interaction chamber along the treated surface only once and in one direction; that is, it is neither pumped back and forth nor repeatedly pumped in a circle along the surface. This results in a simplified device, as back-pumping is no longer necessary, nor is a circulation system. The required low flow velocities are achieved simply, reliably, and, above all, very reproducibly by a pneumatic unit. Furthermore, there are no, or at least significantly fewer, flow fluctuations that would hinder the adhesion of the biomarkers to the conditioned surface.The same advantages arise with the device when it "treats" the surface of the sensor element to condition it, applying conditioning fluid to the surface accordingly. This fluid is also pumped unidirectionally and only once through the interaction chamber.

[0022] The device according to the invention can be designed either only for sample application or only for conditioning the sensor element, or it can be a combined device in which the sensor element is conditioned first and then exposed to the sample of body fluid.

[0023] According to one variant of the invention, the flow rate, or more precisely the volume flow rate, in the interaction chamber is a maximum of 0.4 microliters (µl / min) per minute.

[0024] The pneumatic unit, for example, applies a pressure of 5 to 150 millibar to the liquid in the intake, i.e., relatively low pressure, in order to achieve the low flow velocity.

[0025] To precisely control the low pressure prevailing in the interaction chamber and thus accurately regulate the flow rate, the invention provides for a permanent constriction in the first fluid channel between the intake and the interaction chamber. This constriction causes a pressure drop, allowing the pneumatic unit to operate at a higher pressure that is easier to control.

[0026] The constriction is provided, for example, between the sample intake and the adjacent valve with which the sample is released.

[0027] The flow cross-section at the constriction is a maximum of 40%, in particular a maximum of 15%, of the flow cross-section of the remaining fluid channel, the remaining channel preferably having a constant cross-section.

[0028] The narrowing is, in particular, a conical cone that then widens again conically to avoid turbulence.

[0029] To better control the fluid flow (body fluid or conditioning fluid), especially to define the beginning and end of the flow and to prevent fluid from potentially flowing into other areas of the device than desired, a valve, e.g. a diaphragm valve, is provided in the first fluid channel to open and close it.

[0030] This valve can be controlled by the pneumatic unit, meaning the pneumatic unit combines several functions. It serves both as a pump and as a control device for the valve(s).

[0031] In order to analyze many samples as efficiently as possible in the shortest possible time or to condition surfaces, one variant of the microfluidic device provides for multiple inlets and multiple interaction chambers, with each interaction chamber having its own inlet and its own first and second fluid channel.

[0032] Optionally, only one sensor element can be provided, which closes several or all interaction chambers, i.e., the interaction chambers divided into sections are open on one side, and the sensor element closes all open interaction chambers.

[0033] In particular, each of the recordings has its own valve, so that they can be opened and closed individually.

[0034] The valves are specifically diaphragm valves, which have a common diaphragm clamped between two parts, making the manufacture of the valves cost-effective.

[0035] To save costs, the cleaning of those sections and parts of the device that come into contact with body fluids or previously used conditioning fluid is omitted, as this cleaning process is very complex. For this reason, a disposable fluid guide assembly is provided, which includes at least one interaction chamber, fluid channels or sections thereof, and valves, in particular the aforementioned diaphragm valves. The sensor element is attached to the disposable fluid guide assembly, and the pneumatic unit can be reversibly connected to it. This means, of course, that no fluid may pass from the disposable fluid guide assembly into the pneumatic unit.

[0036] If the aforementioned device according to the invention is designed for the detection of molecular biomarkers in a body fluid, then, of course, the at least one intake is a sample intake for the body fluid. The disposable fluid guide assembly carries the at least one sample intake and / or the at least one disposal container. The at least one sample intake and / or the at least one disposal container is an integral part of the disposable fluid guide assembly or can be directly coupled to it in a fluid-tight manner. In the latter case, the sample intake forms its own closed container, which can then be coupled directly, i.e., without an intermediate tube, to the disposable fluid guide assembly, for example, via a clip or snap connection.

[0037] It should be emphasized that there can be a separate disposal container for each interaction chamber or a common disposal container for several or all interaction chambers.

[0038] Furthermore, a switchable valve can also be provided between the disposal container and the interaction chamber in order to be able to isolate the interaction chamber from the side of the second fluid channel.

[0039] If multiple interaction chambers are provided, the one-way fluid guide assembly can have these interaction chambers, which then have a common sensor element and each have its own first and second fluid channel at its ends.

[0040] The one-way fluid guide assembly, for example, is a component made up of two injection-molded parts, between whose flat surfaces a diaphragm is clamped to form diaphragm valves. The diaphragm separates a fluid side from a pneumatic side. One or both of the opposing flat surfaces have chambers or grooves open towards the diaphragm and closed by the diaphragm, thus forming fluid channels. In the area of ​​the valves, channels on the pneumatic side lead to the pneumatic unit to actuate the diaphragm valves.

[0041] Optionally, at least one inlet for washing fluid, in the form of a washing fluid reservoir, can be provided, which can be connected to the first fluid channel via a valve. The washing fluid is then pumped through the interaction chamber by the pneumatic unit to, for example, give the surface a final cleaning before the body fluid is applied.

[0042] A common washing fluid reservoir can be provided for several or all interaction chambers, or individual receptacles for washing fluid can be used. The washing fluid reservoir(s) are either an integral part of the disposable fluid guide assembly or can optionally be fluid-tightly coupled to it.

[0043] Furthermore, reservoirs for conditioning fluid can be provided. Fluid lines lead from these reservoirs into associated fluid channels, which then connect to the interaction chambers. The pneumatic unit is connected to the reservoirs to pump the conditioning fluid through the interaction chambers.

[0044] If the device is equipped for both conditioning the sensor units and applying a sample, the fluid channels for the conditioning fluid and the fluid channels into which body fluid can be pumped are combined before the interaction chamber. This simplifies manufacturing.

[0045] At least one of the intakes can have a feed opening that is sealed gas-permeable and liquid-tight by a closure with an attached hydrophobic membrane film. This prevents the body fluid or conditioning fluid from leaking out and allows pressure from the pneumatic unit to be introduced into the intake via this membrane film, which seals a corresponding opening in the closure.

[0046] To precisely adjust the low pressure, one or more pressure regulators are advantageous, which detect the pressure either in a receptacle or a fluid channel.

[0047] Furthermore, it can be advantageous to apply a temporary or permanent vacuum to the second fluid channel to draw the fluid out of the interaction chamber and the fluid channels or to assist the flow. It would also be possible to generate pulsation, either in the intake and / or in the second fluid channel, to prevent or clear blockages in the entire flow path.

[0048] The invention also relates to a biomarker detection device with a microfluidic device according to the invention, as previously described. An optical unit is provided which includes an IR source, an IR detector, and a beam path connecting the IR source and the IR detector through the interaction chamber(s). The optical unit is configured to direct the IR radiation generated by the IR source via the beam path through the sensor element to the IR detector. This allows an IR spectrum to be recorded from the surface of the sensor element, thus enabling the identification of any adhering molecular biomarkers.

[0049] Finally, the invention also relates to a method for applying a sample of body fluid to a conditioned sensor element for biomarker detection and / or a method for conditioning a sensor element for biomarker detection, using a microfluidic device according to the invention, as previously mentioned. The method according to the invention comprises the following steps: a) Activating the pneumatic unit and applying pressure to the receiver; b) Opening a valve in the fluid channel; and c) a single passage of a sample of body fluid or conditioning fluid from the first fluid channel through the interaction chamber into the second fluid channel and into the disposal container by means of the applied pressure.

[0050] This means that, even in the inventive method, the sample or conditioning fluid flows only once and unidirectionally through the interaction chamber.

[0051] The aforementioned advantages and individual features in connection with the device can also be used in connection with the method according to the invention.

[0052] The aforementioned pulsing can also be used to dissolve blockages.

[0053] Further features and advantages of the inventions will become apparent from the following description and the accompanying drawings, to which reference is made. The drawings show: - Fig. 1 a schematic sectional view through a biomarker detection device according to the invention in a variant comprising a microfluidic device according to the invention, - Fig. 2 a simplified perspective representation of the biomarker detection device according to Fig. 1, - Fig. 3 a perspective view of a disposable fluid guide assembly used in the biomarker detection device according to one variant, - Fig. 4 a sectional view through a variant of a membrane valve used in the biomarker detection device, - Fig. 5 a sectional view through another variant of a membrane valve used in the biomarker detection device according to the invention, - Fig. 6 a sectional view through a variant showing a container attached to the disposable fluid guide assembly as a receptacle for a liquid, - Fig. 7. A cross-sectional view through an upper section of a container, as a receptacle for a liquid according to a first variant. - Fig. 8 a sectional view through an upper section of a container as a receptacle for a liquid according to a second variant, - Fig. 9 a circuit diagram of a variant of the fluidic device according to the invention as part of the biomarker detection device according to the invention, and - Fig. 10 a perspective transparent view through a section of the lower part of a one-way fluid guide assembly of the microfluidic device in the area of ​​a permanent constriction.

[0054] In Fig. Figure 1 shows a microfluidic device 12 which is suitable for either conditioning a surface of a sensor element 14 for the detection of molecular biomarkers in a body fluid or for wetting the surface with a sample application of body fluid so that biomarkers can adhere to the conditioned surface 16 of the sensor element 14 and be detected.

[0055] One variant provides that the microfluidic device 12 can do both, i.e. first condition the surface 16 and then guide the sample of body fluid along the previously conditioned surface 16.

[0056] For example, protein defects can be detected to diagnose diseases such as those mentioned earlier. The body fluid can, as also mentioned previously, include blood or processed blood.

[0057] If the device 12 is configured to detect biomarkers, it is part of a biomarker detection device 18.

[0058] This biomarker detection device 18 comprises an optical unit 20 with an IR source 22, an IR detector 24, both of which are housed in or attached to an optical housing 26.

[0059] The IR source 22 can be deflected via mirrors 28 to obtain a precisely defined beam path 30. It will be explained later that the beam path 30 is designed such that IR light passes via the mirrors 28 to the surface 16 and from there back, optionally via one or more further mirrors, to the IR detector 24. Due to the interposition of the biomarker-coated surface 16, the beam path 30 has a specific optical spectrum for the biomarkers adhering to the surface 16.

[0060] The microfluidic device 12 comprises a Fig. 3 shown disposable fluid guide assembly 32, which is removed from the device 12 after single use and replaced with a new disposable fluid guide assembly 32.

[0061] The one-way fluid guide assembly 32 comprises, for example (this is not to be understood as a limitation), an upper part 34, which has a flat underside, and a lower part 36 underneath, which also has a flat upper side. A thin elastomeric membrane 38 is clamped between the upper part 34 and the lower part 36; this membrane forms the diaphragm of several diaphragm valves, which will be shown and explained in more detail below.

[0062] The lower part 36, for example, has indentations on its underside, the so-called interaction chambers 40 (see Fig. 1) form. These interaction chambers 40 are closed at the bottom by the common sensor element 14, with the surface 16 facing the interaction chambers 40 and exposed from the interaction chambers 40.

[0063] For each of the multiple interaction chambers 40, a first fluid channel 42, which leads to and opens into the interaction chamber 40, and a second fluid channel 44, which extends from the associated interaction chamber 40 at an end opposite to the opening of the fluid channel 42, are provided. The first fluid channel 42 is thus an inlet and the second fluid channel 44 an outlet for fluid into and out of the associated interaction chamber 40.

[0064] In the illustrated embodiment, at least sections of the two fluid channels 42, 44 are formed in the lower part 36.

[0065] These fluid channels 42 consist of several sections, e.g. one in Fig. 1. A vertically extending section that runs through the lower part 36 in the thickness direction and leads from the top of the lower part 36 to the bottom and to the interaction chamber 40. At the top, there is the adjoining section of the fluid channel, more precisely a top groove, which is covered by the membrane 38 and thus becomes a channel section closed in the cross-sectional direction.

[0066] The first fluid channel 42 leads to a port 46 for a sample of body fluid and / or to a port 48 for conditioning fluid. Fluid channel 42 can also lead to a port 50 for a washing fluid. If fluid channel 42 leads to multiple ports of different fluids, it branches out to extend to the respective ports (chambers, containers, etc.).

[0067] The second fluid channel 44 leads, for example, to a disposal container 52 or to one or more intakes 54 for conditioning fluid already guided along the surface 16.

[0068] The inlets 46, 48, 50, as well as the or several disposal containers 52 and the one or more inlets 54 for conditioning fluid, can either be a single-piece part of the disposable fluid guide assembly 32 or directly or indirectly (via intermediate lines) coupled containers.

[0069] In Fig. Figure 1 shows the image 46 for the sample of body fluid as well as the disposal container 52 as integrated containers, whereas images 48 and 50 as well as image 54 for washing fluid as external containers are shown. However, this is not to be understood as a limitation.

[0070] Fig. Figure 3 shows the illustrated variant of the disposable fluid guide assembly 32, which is shown here in isolation. This assembly features several fluidically separated receptacles 46, which are at least partially depicted as a single block from the outside. Inside the block, however, corresponding to the sample inlet openings 58 shown on the top surface, there are several adjacent chambers for samples of body fluids, each forming an individual receptacle 46 for these samples. In the illustrated embodiment, there are thus eight receptacles 46 for a disposable fluid guide assembly 32, although this is only an example.

[0071] From each of the individual recordings 46, a corresponding, assigned first fluid channel 42 leads to a respective interaction chamber 40 assigned only to this recording 46 and from there via an assigned second fluid channel 44 to a common or one of several disposal containers 52.

[0072] Even in this case Fig. In the variant shown in 3, the disposal containers 52 or the singular disposal container 52 for all intakes 46 are designed as a one-piece molded section of the upper part 34.

[0073] Fig. Figure 6 shows a variant of the design of the receptacles or the container(s), in which either the receptacle 46 or the corresponding container, e.g., the disposal container 52, is designed as a separate part, which is coupled to the upper part 34 via a snap connection 60 (here, for example, snap fingers projecting from the upper part 34). A ring seal 62 ensures a fluid-tight seal.

[0074] To ensure the liquid-filled container is closed upon delivery, it is preferably sealed at its bottom with a film (not shown). A pin protrudes upwards from the upper part 34 in the area of ​​the fluid channel 42, which pierces the film when the receptacle 46 is placed on top and the ring seal 62 has already been compressed somewhat, so that the receptacle 46 is already in close contact with the upper part 34.

[0075] The intake 46 and / or the disposal container 52, as well as, where applicable, the other intakes 48, 50, 54, have a feed opening 58, which is closed by a closure 64 (see Fig. 7) is closed from an elastomeric material, wherein the closure 64 has a hole 66 which is sealed liquid-tight by a hydrophobic membrane film 68. This membrane film 68, however, allows gas exchange, more precisely, air exchange.

[0076] In the variant according to Fig. 7 the hydrophobic membrane film 68 is clamped on the top of the closure 64 between this and an attached pneumatic unit 70.

[0077] The closure 64 has an annular bead 72 on its outer edge, both upwards and downwards, wherein the annular bead 72 are compressed by the axial clamping between the pneumatic unit 70 and the receptacle 46 and / or the disposal container 52 and form ring seals.

[0078] As in Fig. As can also be seen in Figure 7, the attached pneumatic unit has an opening 74 aligned with the hole 66 for supplying air or applying a vacuum.

[0079] The variant according Fig. 8 differs from the one after Fig. 7 by the fact that the closure 64 does not protrude into the feed opening 58, but is placed exclusively on top of the front face of the receptacle 46 or the disposal container 52 and clamps the interposed hydrophobic membrane film 68 between itself and the top in sections.

[0080] Optionally, the pneumatic unit 70 can have one or more annular extensions 76 on the side facing the closure 64 in order to achieve an improved sealing effect around the opening 74.

[0081] In Fig. Not yet mentioned is a holder 77 for the sensor element 14, which surrounds the sensor element 14 and attaches it to the lower part 36. Furthermore, a heated base 78 is provided, which is precisely positioned relative to the optical housing 26 and precisely aligns and holds the disposable fluid guide assembly 32.

[0082] Also visible is the pneumatic unit 70, which is mounted on the top of the upper part 34. The pneumatic unit 70 includes not only the plate-like detail according to the Fig. 1 and Fig. 2, but it is a larger assembly, represented among other things by a pneumatic control 82 in Fig. 1.

[0083] The pneumatic control 82 is, as can be seen in the figure, fluidically coupled to the receptacles 48 and / or 50 for conditioning fluid or washing fluid in order to pressurize these containers and pump the respective fluid into the disposable fluid guide assembly 32.

[0084] Optionally, the pneumatic control 82 can also be coupled with the receptacle 54 for previously used conditioning fluid in such a way that a negative pressure is created in the corresponding containers. The channels or lines extend from the pneumatic control 82 into Fig. 1 shown with broken lines, in Fig. 2. They have been omitted for the sake of clarity.

[0085] However, in Fig. 2 to recognize that the recordings 48, 50, 54 are coupled to a control block 84, 86, which in turn optionally carries valves 88, 91, which are coupled to the channels or lines to open or close them.

[0086] For the detachable coupling and decoupling of the pneumatic unit 70 to the respective disposable fluid guide assembly 32, a mounting unit 80 can be provided, which is Fig. 2 is shown.

[0087] Furthermore, an electrical control unit 90 is shown, which is coupled via signal lines 92 to valves and the pneumatic unit 70 together with its pneumatic control unit 82.

[0088] Finally, in Fig. Figure 2 also shows a volume flow measuring unit 95, which is shown by way of example upstream of the recording 54 after the disposable fluid guide assembly 32, but can also be positioned at any other point in the first or the second fluid channel 42, 44. It serves to control the flow velocity in the fluid channels 42, 44.

[0089] In Fig. Figure 9 shows a fluidic circuit diagram of the biomarker detection device 18, more precisely its microfluidic device 12.

[0090] As previously explained, there is the central disposable fluid guide assembly 32 with the sensor element 14, which closes several interaction chambers 40 at their open side.

[0091] The interaction chambers 40, of which only two are shown with reference symbols for clarity, are each connected to the associated first fluid channels 42 and connected to the associated second fluid channels 44.

[0092] The receptacles 46 for accommodating samples of body fluids, for example from different patients, are located on the disposable fluid guidance assembly 32 or are an integral part thereof.

[0093] In the fluid channel 42 leading from the respective recording 46 to the associated interaction chamber there is a valve 94, in the form of a diaphragm valve, to start or stop the supply of body fluid to the interaction chamber 40.

[0094] Each fluid channel 42 optionally splits and also leads to the intake 50 for washing liquid, wherein a valve 96 in the form of a diaphragm valve is also located in the corresponding channel section in order to individually supply washing liquid to each fluid channel 42, which then leads the washing liquid to the interaction chamber 40.

[0095] Fig. Figure 9 optionally shows that a common receptacle 50 for washing fluid can be provided for all interaction chambers 40 and that this receptacle 50 can also be located on the disposable fluid guide assembly 32 or be an integral part of it.

[0096] The inlets 48 for conditioning fluid are, for example, arranged outside the disposable fluid guide assembly 32, with several different conditioning fluids optionally provided, each with its own container and which can be supplied to the upstream end of the fluid channel 42 via its own valve 98. Different conditioning fluids can be alternately fed through the corresponding interaction chamber 40 via the valves 98.

[0097] The second fluid channels 44 lead to a central disposal container 52, where optional valves 100 are also provided in each fluid channel 44 immediately upstream of the disposal container 52.

[0098] Each fluid channel 44 can branch and lead via another valve 102, also a diaphragm valve, to one or more receptacles 54 for used conditioning fluid. It may be advantageous to have a separate receptacle 54 for each type of conditioning fluid, in order to reuse this relatively pure fluid or dispose of it separately.

[0099] Alternatively, the inlets 54 can be omitted, so that the fluid channels 44 terminate in the area of ​​the disposal container 52. In this case, the volumetric flow measuring unit 95 should measure upstream of the disposal container 52, either in fluid channel 44 or in fluid channel 42.

[0100] The pneumatic unit 70 including pneumatic control 82, which is in Fig. 9, which are represented as a block, comprises a first pneumatic pump 106, a second pneumatic pump 108 and several electrically controllable valves 110, of which only one is symbolically provided with a reference sign.

[0101] A vacuum line 112 is fluidically coupled to the pump 106, and a pressure line 114 and a drive line 116 are fluidically coupled to the second pump 108.

[0102] From the pneumatic unit 70, lines 118 lead to the inlets 48, a line 120 to the inlet 50, individual lines 122 to the inlets 46, a line 125 to the disposal container 52, and, if necessary, one or more lines 126, 128 to the inlets 54.

[0103] By switching corresponding valves 110 in the pneumatic unit 70, the receptacles 48 can be pressurized via the lines 118, the receptacle 50 via the line 120 and the receptacles 46 for body fluid via the lines 122.

[0104] Optionally, a vacuum can be applied to the disposal container 52 via line 125, and similarly to the inlets 54 for conditioning fluid via lines 126 and 128. The vacuum can optionally be pulsed to prevent or clear blockages in the fluid channels 42 and 44.

[0105] For clarity, lines 130 to the individual valves 94, 96, 100, 102 are only indicated. These lines 130 are pneumatic lines with which the aforementioned valves 94, 96, 100, 102, which are designed as diaphragm valves, can be controlled either individually or in groups (for example, all valves 94 together).

[0106] Before explaining the operation of the biomarker detection device 18 and the fluidic device 12, the previously mentioned valves, designed as diaphragm valves, will be described in the Fig. 4 and Fig. 5 briefly introduced.

[0107] As mentioned, a thin elastomeric membrane 38 (see Fig. 4) clamped between the upper part 34 and the lower part 36. The diaphragm 38 fluidically separates the pneumatic side in the upper part 34 from the fluid side in the lower part 36.

[0108] In the upper part 34, chambers 121 are provided which are opposite the valve seats 123 on the fluid side in order to optionally press the diaphragm 38 against the valve seat 123 or to lift or allow the diaphragm 38 to be lifted in this area.

[0109] The fluid side has corresponding sections of fluid channels 42, 44 extending in the direction of the thickness of the lower part 36 and, beyond the respective valve seat 123, sections 124 that form parts of a groove on the top of the lower part 36. These grooves are sections of the respective fluid channels 42, 44 that are closed circumferentially by the diaphragm 38.

[0110] To precisely position and seal the membrane 38, it can have a protruding bead 126 on one side (see Fig. 4), which can, for example, be received in a corresponding groove 128 in one of the parts, here in the upper part 34. The bead 126 can, for example, extend around each valve and seal this section of the diaphragm 38 against an adjacent section that forms another valve.

[0111] Fig. Figure 5 shows a membrane 38 which has a projecting bead 126 on each of its opposite sides, which then projects into a corresponding groove 128 in the upper part 34 and in the lower part 36.

[0112] Optional, this is not dependent on the embodiment according to Fig. 5 limited, the diaphragm 38 in the area of ​​the valve seat 123 can also be made thinner.

[0113] As in Fig. Figure 3 shows the receptacle 46 for the sample of body fluid on the upper part 34, and the pneumatic unit 70 is coupled to the upper part 34 from above, so that pressure can be applied inside the receptacle 46 via the hydrophobic membrane film 68.

[0114] Since the pneumatic side is formed in the upper part 34 and the hydraulic side in the lower part 36, a corresponding channel must lead from the receptacle 46 through a permanent opening in the diaphragm 38 into the lower part 36, where the remaining part of the fluid channel 42 and the fluid channel 44 are then formed.

[0115] Fig. Figure 10 shows an x-ray-like view into the lower part 36 and on its upper side with the membrane 38 removed.

[0116] The lower part 36 has a trough-like depression 140, from the bottom of which a central opening 142 leads into an outlet pipe 144, which runs towards the underside of the lower part 36.

[0117] A throttle channel 146 connects to the outlet pipe 144 and extends to a pipe 148, which then runs to the top of the lower part 36 and terminates there in the valve seat 123. Section 124 in Fig. 4 is therefore a groove that is in Fig. 10 can be seen and is then closed by the membrane 38.

[0118] Also in Fig. Figure 10 shows section 150 of the fluid channel 42, which extends from inlet 50 or inlet 48 and merges with line 148 at the valve seat 123 when the valve is open. Section 150, line 148, throttle channel 146, and outlet line 144 form sections of the fluid channel 42.

[0119] The throttle channel 146 itself, or a section thereof, forms a permanent constriction 152 of the fluid channel 42 upstream of the interaction chamber 40. For example, the constriction 152 is as shown in Fig. 9 can be seen, located between recording 46 and valve 94.

[0120] The constriction 152 can be achieved, for example, by a plate element 154, which is screwed from below onto the lower part 36 in the area of ​​the throttle channel 146, which would otherwise be open at the bottom. This narrows the cross-section of the throttle channel 146 in the area of ​​the constriction 152 to a slot-like cross-section. The plate element 154 can, for example, project into the throttle channel 146 with a curved upper surface.

[0121] The flow cross-section in the area of ​​the constriction 152 is a maximum of 40%, in particular a maximum of 15% of the flow cross-section of the remaining fluid channel 42.

[0122] The following explains the operation of the microfluidic device 12.

[0123] In one variant, the device 12 is intended solely for conditioning the sensor element 14 and optionally does not need to have the receptacles 46 and 50. It would also suffice to either omit the disposal container 52 or the receptacles 54.

[0124] By applying pressure, conditioning fluid from the receptacles 48 enters the first fluid channels 42 through the opening of the respective valves 98 and flows along the respective interaction chambers 40 to treat the surface of the sensor unit. Different conditioning fluids can be pumped alternately from the respective assigned receptacle 48 into the interaction chambers 40.

[0125] The conditioning fluid flows through the interaction chambers 40 only once and in one direction, so the pressure in the recordings must be very low. However, due to the pressure drop at the constriction 152, it can be higher than it would be without such a constriction 152.

[0126] The conditioning fluid then flows either into the respective container in the assigned receptacle 54 or into the common disposal container 52, where a vacuum may be applied.

[0127] It may then be necessary to flush the fluid channels 42 and the interaction chambers 40 by applying pressure to the intake 50 for washing fluid and opening the valves 96. This washing fluid then enters the disposal container 52 through the open valves 100 and the closed valves 102.

[0128] Finally, the samples of body fluid from the recordings 46 are pumped into the fluid channels 42 by applying pressure. The body fluid also flows unidirectionally and only once through the respective interaction chamber 40 and then reaches the disposal container 52.

[0129] The device can be used either solely for conditioning or solely for applying body fluid samples and detecting molecular biomarkers such as protein defects. In the latter case, the corresponding images 48, possibly 50, and 54 are omitted.

[0130] However, the device can also be used for both purposes, as shown in Fig. 9 shown.

[0131] For the detection of biomarkers, the device 12 is part of the in Fig. 1 Biomarker detection device 18 shown, in which the beam path 30 is ultimately directed via the IR source 22 to the sensor element 14 in order to record an IR spectrum from the surface 16 and thus to obtain in the control a conclusion about the biomarker adhering to the conditioned surface 16.

[0132] The flow rate of the conditioning fluid and the sample of body fluid is a maximum of 0.4 mm / min in the interaction chamber 40.

[0133] The pneumatic unit 70 only applies a pressure of 5 to 150 mbar to the liquid in the respective intake. QUOTES INCLUDED IN THE DESCRIPTION

[0000] This list of documents cited by the applicant was automatically generated and is included solely for the reader's convenience. The list is not part of the German patent or utility model application. The DPMA accepts no liability for any errors or omissions. Cited patent literature

[0000] US 2020 / 0 141 866 A1

[0009]

Claims

[1] Microfluidic device (12) for applying a sample of body fluid to a conditioned sensor element for the detection of molecular biomarkers in the body fluid and / or for conditioning the sensor element (14) by means of a conditioning fluid, comprising at least one interaction chamber (40) which is partially closed by a sensor element (14), wherein a first fluid channel (42) opens at one end of the at least one interaction chamber (40) and a second fluid channel (44) opens at the opposite end, at least one first inlet (46) for receiving the body fluid or the conditioning fluid, which is fluidically connected to the first fluid channel (42) at an end of the at least one interaction chamber (40) forming a fluid inlet, wherein the second fluid channel (44) leads from its associated interaction chamber (40) to a disposal container (52), wherein the sensor element has a sensor surface facing the interaction chamber, to which biomarkers adhere in the conditioned state, and comprising a pneumatic unit (70) connected to the at least one first intake (46) and configured to pressurize it and pump the body fluid or conditioning fluid once and unidirectionally through the at least one interaction chamber (40) along the treated surface (16) and subsequently into the at least one disposal container (52), so that, in the case of body fluid, biomarkers adhere to the surface (16) or, in the case of conditioning fluid, the surface is conditioned. [2] Microfluidic device (12) according to claim 1, characterized by , that at least one fluid channel (42) between the intake (46) and the interaction chamber (40) has a permanent narrowing (152). [3] Microfluidic device (12) according to claim 2, characterized by, that the flow cross-section at the constriction (152) is a maximum of 40%, in particular a maximum of 15%, of the flow cross-section of the remaining fluid channel (42). [4] Microfluidic device (12) according to any one of the preceding claims, characterized by , that at least one valve (94, 96) is provided in the first fluid channel (42) to open and close it. [5] Microfluidic device (12) according to any one of the preceding claims, characterized by , that several inlets (46, 48) and several interaction chambers (40) are provided, with each interaction chamber (40) having its own inlet (46, 48) and its own first and second fluid channel (44). [6] Microfluidic device (12) according to claims 4 and 5, characterized by , that each intake (46, 48) is assigned its own valve (94, 96). [7] Microfluidic device (12) according to claim 6, characterized by, that the valves (94, 96) are diaphragm valves which have a common diaphragm (38) clamped between two parts. [8] Microfluidic device (12) according to any one of the preceding claims, characterized by , that a one-way fluid guide assembly (32) is provided which has at least one interaction chamber (40), fluid channels (42, 44) or sections of the fluid channels (42, 44) and valves (94, 96, 100, 102), in particular diaphragm valves, wherein the sensor element (14) is attached to the one-way fluid guide assembly (32) and the pneumatic unit (70) is reversibly coupled to the one-way fluid guide assembly (32). [9] Microfluidic device (12) according to claim 8 for the detection of molecular biomarkers in a body fluid, characterized by, that the at least one intake (46) is a sample intake for the body fluid, that the disposable fluid guide assembly (32) carries the at least one sample intake and / or the at least one disposal container (52), and that the at least one sample intake and / or the at least one disposal container (52) is an integral part of the disposable fluid guide assembly (32) or is directly coupled to it in a fluid-tight manner. [10] Microfluidic device (12) according to claim 8 or 9, characterized by , that the disposable fluid guide assembly (32) has several interaction chambers (40) which have a common sensor element (14) and which each have their own first and second fluid channels (42, 44) at their ends. [11] Microfluidic device (12) according to any one of the preceding claims, characterized by, that at least one receptacle (50) designed as a washing liquid container for washing liquid can be connected to the first fluid channel (42) via a valve (96) in order to convey washing liquid through the interaction chamber (40). [12] Microfluidic device (12) according to one of claims 8 to 10 and additionally according to claim 11, characterized by , that a common washing fluid reservoir is provided for several interaction chambers (40), which is an integral part of the disposable fluid guide assembly (32) or is fluid-tightly coupled to it. [13] Microfluidic device (12) according to any one of the preceding claims, characterized by , that at least one receptacle (46, 50) has a feed opening (58) which is sealed gas-permeable and liquid-tight by a closure (64) with a hydrophobic membrane film (68) attached thereto, and wherein the pneumatic unit (70) introduces pressure into the receptacle (46, 50) via the membrane film (68). [14] Biomarker detection device (18) with a microfluidic device (12) according to one of claims 1 to 13, wherein the biomarker detection device (18) comprises an optical unit (20) having an IR source (22), an IR detector (24) and a beam path (30) connecting the IR source (22) and the IR detector (24) through the or several interaction chambers (40), wherein the optical unit (20) is configured to direct the IR radiation generated by the IR source (22) via the beam path (30) over the sensor element (14) to the IR detector (24) in order to record an IR spectrum from the surface (16) of the sensor element (14). [15] Method for applying a sample of body fluid to a conditioned sensor element (14) for biomarker detection and / or for conditioning a sensor element (14) for biomarker detection, using a microfluidic device (12) according to any one of claims 1 to 13, wherein the method comprises the following steps: a) Activating the pneumatic unit (70) and applying pressure to the receiver (46, 48); b) Opening a valve (94, 98) in the fluid channel (42); and c) a single passage of a sample of body fluid or conditioning fluid from the first fluid channel (42) through the interaction chamber (40) into the second fluid channel (44) and into the disposal container (52) by the applied pressure.

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