DEVICE AND METHOD FOR ANALYZING BIOLOGICAL SAMPLES

Abstract

The invention relates to a device (1) for analyzing biological samples (S) comprising a substrate (2) for receiving a biological sample (S), wherein the substrate (2) comprises or consists of a fibrous material (F) configured to form a stationary phase which retains molecules in a biological sample (S) dissolved in a liquid mobile phase depending on molecular weight and/or polarity of the molecules, a first electrode (41) and a second electrode (42), which are arranged along a first axis (A1) and configured to generate an electric field acting along the first axis (A1) when an electric potential difference is provided between the first electrode (41) and the second electrode (42), so that charged molecules contained in the biological sample (S) are movable through the substrate (2) along the first axis (A1) and/or separable by their molecular weight, their polarity and/or their charge, wherein the substrate (2) comprises a chemical lysing agent (L) capable of lysing the biological sample (S). Furthermore, a method for analyzing biological samples (S) using the device (1) is provided.

Claims

1. A device (1) for analyzing biological samples (S) comprising a substrate (2) for receiving a biological sample (S), wherein the substrate (2) comprises or consists of a fibrous material (F) configured to form a stationary phase which is capable of retaining molecules in a biological sample (S) dissolved in a liquid mobile phase depending on molecular weight and/or polarity of the molecules, a first electrode (41) and a second electrode (42), which are arranged along a first axis (A1) and configured to generate an electric field acting along the first axis (A1) when an electric potential difference is provided between the first electrode (41) and the second electrode (42), so that charged molecules contained in the biological sample (S) are movable through the substrate (2) along the first axis (A1) and/or separable by their molecular weight, their polarity and/or their charge, wherein the substrate (2) comprises a chemical lysing agent (L) capable of lysing the biological sample (S), characterized in that the device (1) comprises a third electrode (43), wherein the second electrode (42) and the third electrode (43) are arranged along a second axis (A2) which is non-parallel to the first axis (A1) and configured to generate an electric field acting along the second axis (A2) when an electric potential difference is applied between the second electrode (42) and the third electrode (43).

2. The device (1) according to claim 1, characterized in that the lysing agent (L) is disposed within the substrate (2) in dry form, wherein the lysing agent (L) is dissolvable upon applying a liquid biological sample (S) to the substrate (2), such that the biological sample (S) is lysed.

3. The device (1) according to claim 1, characterized in that the substrate (2) comprises a barrier structure (B) which is impermeable to the liquid mobile phase, wherein the barrier structure (B) forms at least one channel in the substrate (2), is the channel being permeable to the liquid mobile phase, and the channel being limited by the barrier structure (B).

4. The device (1) according to claim 1, characterized in that the first electrode (41), the second electrode (42) and/or the third electrode (43) is embedded into the fibrous material (F) of the substrate (2) or connected to the fibrous material (F) of the substrate (2), particularly wherein the first electrode (41), the second electrode (42) and/or the third electrode (43) is printed onto the substrate (2).

5. The device (1) according to claim 1, characterized in that the second electrode (42) comprises a cross-sectional area perpendicular to the first axis (A1), which is smaller than a cross-sectional area perpendicular to the first axis (A1) of the first electrode (41), particularly so that charged molecules in the biological sample (S) are focused in the substrate (2) while moving through the electric field between the first electrode (41) and the second electrode (42).

6. The device (1) according to claim 1, characterized in that the first electrode (41) forms a grid structure (41b).

7. The device (1) according to claim 1, characterized in that a DNA polymerase and PCR primers are disposed in the substrate (2), so that a DNA sequence of the biological sample (S) can be amplified by PCR by means of the DNA polymerase if the PCR primers contain complementary DNA sequences to said DNA sequence.

8. The device (1) according to claim 1, characterized in that the device (1) comprises a polymer gel (H), particularly a hydrogel, in contact with the substrate (2), so that liquid is exchangeable between the polymer gel (H) and the substrate (2), particularly wherein the polymer gel (H) is switchable by setting a physical parameter, particularly changing a temperature, a pH, or an ionic strength, providing electromagnetic radiation, or initiating a chemical reaction, so that liquid is exchanged between the polymer gel (H) and the substrate (2).

9. The device (1) according to claim 1, characterized in that the device (1) comprises a heating element (70) for heating, particularly periodic heating, of the substrate (2), particularly wherein the heating element (70) is formed as a heating electrode, wherein the heating electrode is connected to an outside of the device (1) via electrically conductive contact tabs (71), wherein ends (72) of the first and second contact tabs (71) are configured to be electrically connected to the poles of a voltage source, so that an electric current flows through the heating electrode thereby heating the substrate (2) by its electric resistance.

10. The device (1) according to claim 1, characterized in that the substrate (2) comprises a marker for labeling molecules contained in the biological sample, particularly wherein the marker comprises a dye or wherein the marker is detectable by its redox properties.

11. The device (1) according to claim 1, characterized in that the device is at least partially transparent, so that the molecules contained in the biological sample (S) can be analyzed by an optical detector (80), particularly a camera, more particularly a camera contained in a handheld device such as a smart phone or a tablet computer.

12. The device (1) according to claim 1, characterized in that the device (1) comprises a plurality of layers (10, 20, 30) comprising said fibrous material (F), wherein the layers (10, 20, 30) are arranged on top of each other, so that the fibrous material (F) of the layers (10, 20, 30) forms the substrate (2), wherein the layers (10, 20, 30) extend in respective planes which are non-parallel to the first axis (A1), so that charged molecules contained in the biological sample (S) are movable through the layers (10, 20, 30) by means of the electric field acting along the first axis (A1) between the first electrode (41) and the second electrode (42), particularly wherein the planes in which the layers (10, 20, 30) extend, are parallel to the second axis (A2), so that the charged molecules contained in the biological sample (S) are movable in the respective layer (10, 20, 30) and separable by their molecular weight, polarity and/or charge.

13. The device (1) according to claim 12, characterized in that the device (1) comprises a first layer (10), wherein the fibrous material (F) of the first layer (10) comprises the lysing agent (L), a second layer (20) in contact with the first layer (10), wherein the second layer (20) is configured to filtrate the biological sample (S) when the biological sample (S) passes through the second layer (20), and a third layer (30) in contact with the second layer (20), wherein the third layer (30) comprises said second electrode (42) and said third electrode (43), such that molecules in said biological sample (S) are separable by molecular weight, polarity and/or charge in the third layer (30) using the electric field acting between the second electrode (42) and the third electrode (43), wherein particularly the fibrous material (F) of the third layer (30) comprises said marker and/or said DNA polymerase and said PCR primers, particularly wherein the fibrous material (F) of the third layer (30) is in contact with said heating element (70), particularly wherein said polymer gel (H) is arranged in said first layer (10) and/or said second layer (20).

14. A method for analyzing biological samples (S) by means of the device (1) according to claim 10, wherein a. a biological sample (S) is applied to the substrate (2), such that the biological sample (S) is in contact with said lysing agent (L), resulting in lysis of the biological sample (S); b. an electric potential difference is provided between the first electrode (41) and the second electrode (42), such that the biological sample (S) moves through the substrate (2) and is separated by its molecular weight, polarity and/or charge; c. optionally, the substrate (2) is periodically heated, such that a DNA sequence of the biological sample (S) is amplified by PCR in the substrate (2); d. optionally, an electric potential difference is provided between the second electrode (42) and the third electrode (43), such that the biological sample (S) is separated by its molecular weight, polarity and/or charge, particularly wherein the current or the voltage between the second electrode (42) and the third electrode (43) is measured over time, wherein properties of the biological sample (S) are derived from the measured current or voltage; and e. optionally, the biological sample (S) is analyzed by detecting the marker in the substrate (2).

Description

[0150] The invention is further illustrated by the following examples and figures, from which further embodiments and advantages can be drawn. These examples are meant to illustrate the invention but not to limit its scope.

[0151] FIG. 1 shows a perspective exploded view of a device according to a first embodiment of the invention;

[0152] FIG. 2 shows a part of the device depicted in FIG. 1 illustrating details of the first layer, the second layer and the third layer;

[0153] FIGS. 3 to 6 illustrate different modes of operation of the device according to the first embodiment of the invention;

[0154] FIG. 7 illustrates optical detection of molecules in the biological sample on the substrate of the device according to the invention;

[0155] FIG. 8 shows a further embodiment of the device according to the invention comprising a lid including the first electrode;

[0156] FIG. 9 shows a further embodiment of the device according to the invention comprising resistance heating elements disposed on a cover layer;

[0157] FIG. 10 shows a further embodiment of the device according to the invention comprising resistance heating elements disposed on fourth layer,

[0158] FIG. 11 shows a further embodiment of the device according to the invention realizing a diagonal sample path through the substrate layers between the first electrode and the second electrode;

[0159] FIG. 1 shows a device 1 according to the present invention comprising a plurality of layers stacked along a first axis A1. The device 1 comprises from top to bottom according to the view of FIG. 1: a cover 60, a cover layer 50, a first layer 10, a second layer 20 and a third layer 30, each formed from the fibrous material F with an additional barrier structure B, and a further cover layer 50.

[0160] In the example described here, the liquid mobile phase to be used with the device 1 is particularly aqueous. Consequently, the fibrous material F is water-permeable, wherein the fibrous material F is particularly paper. In addition, the cover 60, the cover layers 50, and the barrier structure B is particularly water-impermeable. Of course, if a hydrophobic liquid mobile phase (e.g. based on an organic solvent) is to be used with the device 1, the fibrous material F is particularly hydrophobic (and therefore permeable to the hydrophobic liquid mobile phase), and the cover 60, the cover layers 50 and the barrier structure B are particularly hydrophilic or water-permeable (and therefore impermeable to the hydrophobic liquid mobile phase).

[0161] The cover 60 and the two cover layers 50 protect the first layer 10, the second layer 10, and the third layer 30 from ingress of water and contaminations, particularly during transport and storage of the device 1. The upper cover layer 50 comprises an opening 51 (particularly having a rectangular shape) through which the biological sample S may be applied to the first layer 10 after the cover 60 has been removed.

[0162] In the embodiment depicted in FIG. 1, the first layer 10 comprises a first area of fibrous material F (particularly having a rectangular shape) surrounded by the barrier structure B (the barrier structure B is displayed crosshatched). This fibrous material F forms a part of the substrate 2 as specified below. When the upper cover layer 50 is placed on top of the first layer 10, the opening 51 of the upper cover layer 50 is congruent with the first area of accessible (i.e., not being covered by the barrier structure B) fibrous material F of the first layer 10.

[0163] FIG. 1 further displays heating elements 70 positioned below the lower cover layer 50. During operation of the device 1, the lower cover layer 50 is particularly in direct contact with the lower cover layer 50, and the lower cover layer 50 is in direct contact with the third layer 30, such that the third layer 30 may be heated by the heating elements 70.

[0164] FIG. 2 shows details of the first layer 10, the second layer 20 and the third layer 30 of the device 1, omitting the cover layers 50, cover 60 and heating elements 70 for clarity.

[0165] As shown in FIG. 2, the first area 2a of fibrous material F of the first layer 10 comprises a chemical lysing agent L which is particularly embedded into and essentially equally distributed throughout the fibrous material F of the first layer 10. The lysing agent L is capable of lysing cells (bacteria, archaea or eukaryotes) contained in the biological sample S, e.g. by denaturation of lipid membranes and/or degradation of proteins which are part of the cell structure. In particular, the lysing agent L is disposed in the fibrous material F in dry or lyophilized form, such that the lysing agent L is dissolved when liquid is added, e.g. by application of the liquid biological sample S or by release of the liquid from a switchable polymer gel H (see below).

[0166] The first layer 10 further comprises a first electrode 41 shaped as grid structure 41b which comprises parallel rods or bars 41c from an electrically conductive material extending in the plane of the first layer 10. The bars 41c are surrounded by a frame 41d (particularly having a rectangular shape) arranged around the first area 2a of accessible fibrous material F. A contact tab 41a, particularly from the same electrically conductive material, connects the electrode 41, particularly the frame 41d, to an edge of the first layer 10, such that the contact tab 41 can be electrically connected to a positive or negative pole of an external voltage source. The electrode 41 is particularly printed onto the fibrous material F of the first layer 10, i.e. by an electrically conductive ink.

[0167] Furthermore, the first layer 10 comprises a circular opening 11 harboring a switchable polymer gel H. The opening 11 is separated from the first area 2a of accessible fibrous material F forming part of the substrate 2 by the barrier structure B, such that liquid released from the polymer gel H cannot access the substrate 2 via the first layer 10.

[0168] Similar to the first layer 10, the second layer 20 comprises a second area 2b of uncoated, accessible fibrous material F (particularly having a rectangular shape) surrounded by the barrier structure B, the uncoated fibrous material F of the second area 2b forming a part of the substrate 2. Said second area 2b of the second layer 20 is congruent with the first area 2a of the first layer 10 when the first layer 10 and the second layer 20 are placed on top of each other. The second layer 20 also comprises a circular opening 21 which is congruent with the opening 11 of the first layer 10 when the first layer 10 and the second layer 20 are placed on top of each other. The opening 21 may harbor a further polymer gel H, or the openings 11, 21 of the first layer 10 and the second layer 20 may jointly harbor a polymer gel H.

[0169] The third layer 30 comprises a third area 2c of freely accessible (i.e. uncovered by the barrier structure B) fibrous material F (particularly having a rectangular shape) surrounded by the barrier structure B. The third area 2c is larger (extends further to the right in FIG. 2) than the first area 2a of the first layer 10 and the second area 2b of the second layer 20. When the first layer 10, the second layer 20 and the third layer 30 are stacked on top of each other, the third area 2c overlaps with the first area 2a of the first layer 10 and the second area 2b of the second layer 20, such that the three areas 2a, 2b, 2c of fibrous material F jointly form the substrate 2 providing a channel through which the biological sample S may migrate as specified below.

[0170] The fibrous material F of the third area contains PCR reagents P (i.e. a DNA polymerase, PCR primers and deoxy ribonucleotides), which are particularly provided in dry or lyophilized form and are particularly equally distributed in the fibrous material F of the third layer 30. Thereby, if a liquid biological sample S containing DNA or RNA with sequences complementary to the PCR primers provided in the fibrous material enters the fibrous material F of the third layer 30, and if the third area 2c of fibrous material F is periodically heated and cooled to appropriate melting, annealing and elongation temperatures, the complementary DNA or RNA sequence may be amplified by PCR within the fibrous substrate F.

[0171] Furthermore, the third layer 30 comprises a second electrode 42 and a third electrode 43 which are arranged on opposite edges of the third area 2c and connected to respective outer edges of the third layer 30 by contact tabs 42a, 43a, such that the second electrode 42 and the third electrode 43 may be electrically connected to poles (particularly opposite poles of an external voltage source via the contact tabs 42a, 43a.

[0172] Moreover, when the first layer 10, the second layer 20 and the third layer 30 are placed on top of each other, the polymer gel H or polymer gels H disposed in the openings 11, 21 of the first layer 10 and the second layer 20 are in contact with the third area 2c of the third layer 30, such that liquid may be released from the polymer gel(s) H into the fibrous material F of the third layer 30, and/or such that liquid may be withdrawn from the fibrous material F of the third layer 30 into the polymer gel(s) H, as further specified below.

[0173] In particular, to manufacture the layers 10, 20, 30, thin sheets of the fibrous material F are provided and optionally cut into an appropriate size. Subsequently, the layers are particularly coated with a hydrophobic material, such as wax, to form the barrier structure B, and the openings 11, 21 of the first and second layer 10, 20 are particularly cut out. The first electrode 41, the second electrode 42 and the third electrode 43 may then be printed onto the first layer 10 and the third layer 30 using an electrically conductive ink. Finally, the lysing agent L may be applied to the first layer 10, particularly by applying a liquid solution containing the lysing agent L onto the first rectangular area 2a, and subsequently drying the first layer 10. Likewise, the PCR reagents P may be applied onto the third area 2c of the third layer as a liquid solution and subsequently dried. The layers 10, 20, 30 may then be stacked above each other, and the polymer gel H may be applied in the openings 11, 21 of the first layer 10 and/or second layer 20. Finally, the cover layers 50 and cover 60 may be placed on the stack as depicted in FIG. 1.

[0174] The use of the device 1 is described hereafter with reference to FIGS. 3-6.

[0175] As shown in FIG. 3, a liquid biological sample S is first applied to the first area 2a of the first layer 10, e.g. by manually or automatically pipetting the sample S onto the fibrous material F of the first area 2a. Thereby, in particular, the dry lysing agent L disposed in the fibrous material F is dissolved in the liquid biological sample S and cells in the biological sample S are lysed by the lysing agent L, thus releasing biomolecules such as DNA, RNA and proteins from these cells.

[0176] Since the first area 2a of fibrous material F in the first layer 10 is in contact with the second area 2b of the second layer 20 and the third area 2c of the third layer 30, the lysed liquid biological sample S is subsequently drawn into the fibrous material F of the second area 2b and the third area 2c by capillary forces. Therein, the liquid, in which the biological sample S is dissolved, serves as a mobile phase and the fibrous material F of the layers 10, 20, 30 serves as a stationary phase, whereby the fibrous material F, particularly the second area 2b of the second layer 20, filtrates the biological sample S by separating the components of the biological sample S by molecular weight due to the chromatographic effect. In this manner, in particular, larger particles, such as cell debris, may be separated from the biomolecules (i.e., DNA, RNA and proteins) of the biological sample, and particularly remain in the fibrous material F of the second layer 20. Optionally, the fibrous material F may also be adapted to separate the biological sample by polarity, depending on the hydrophobicity of the fibrous material F and the solvent of the biological sample S.

[0177] To facilitate the movement of the biological sample from the first layer 10 to the second layer 20 and the third layer 30, polymer gels H capable of swelling (taking up liquid) may be placed in the openings 11, 21 of the first and second layer 10, 20 to draw the solvent of the biological sample through the first layer 10, the second layer 20 and the third layer 30.

[0178] Alternatively or additionally, the device 1 may be placed in a liquid reservoir serving as a mobile phase for chromatography and/or as electrophoresis buffer.

[0179] As illustrated in FIG. 4, an electric field may be provided between the first electrode 41 in the first layer 10 and the second electrode 42 in the third layer 30, particularly by connecting the contact tabs 41a, 42a to opposite poles of an external voltage source to electrophoretically move components of the biological sample S from the first layer 10 to the second layer 20 and the third layer 30. In particular, a negative potential may be applied to the first electrode 41 and a positive potential may be applied to the second electrode 42 as depicted in FIG. 4. In particular when the biological sample S is dissolved in a buffer having a pH range between about 6 and about 8, most biomolecules (DNA, RNA and proteins) in the biological sample S are negatively charged. Therefore, most biomolecules will electrophoretically migrate toward the second electrode 42 in the third layer 30 when the first electrode 41 has a negative potential and the second electrode 42 has a positive potential.

[0180] In the setup depicted in FIG. 3-6, the first electrode 41 is a grid electrode stretching over the entire first area 2a of the first layer 10, and the second electrode 42 is formed as an elongated rectangular tab having a smaller cross-sectional area perpendicular to the first axis A1 than the first electrode 41.

[0181] Thus, as shown in FIG. 4, the biological sample B is focused while passing through the first layer 10 and the second layer 20 to the third layer 30, as indicated by the sample path SP in FIG. 4. In other words, as a result of the convergence of the electric field due to the smaller second electrode 42, the molecules of the biological sample are concentrated to a smaller partial area of the third layer 30.

[0182] In particular, as illustrated in FIG. 5, the third layer 30 containing the PCR reagents P and the biological sample S is subsequently periodically heated (and optionally cooled) by the heating elements 70 below the third layer 30 to perform a PCR amplification of DNA or RNA contained in the biological sample S (heating of the PCR reagents 70a). To ensure that the third layer 30 does not dry out during the heating cycles of the PCR reaction, the polymer gel H arranged in the opening 11, 21 of the first layer and/or second layer may be activated to release liquid into the third layer 30, in particular by heating with the heating elements 70 (heating of the switchable polymer gel 70b, in case the polymer gel H is temperature-switchable). Of course, the PCR step may be omitted, in particular if analysis of other biomolecules than nucleic acids (e.g. proteins) contained in the biological sample S is desired.

[0183] Finally, as shown in FIG. 6, the (particularly previously amplified) molecules contained in the biological sample S may be separated by electrophoresis in the third layer 30. To this end, an electric potential difference is applied between the second electrode 42 and the third electrode 43, such that an electric field is generated in the plane of the third layer 30. In particular, the second electrode 42 may be connected to a negative pole 92 of a voltage source and the third electrode 43 may be connected to a positive pole 91 of the voltage source, such that predominantly negatively charged biomolecules (such as DNA, RNA and proteins) migrate from the second electrode 42 to the third electrode 43.

[0184] The voltage source may be configured to provide a constant voltage between the second electrode 42 and the third electrode 43 or a constant current through the third layer 30. During electrophoresis, the current in the third layer 30 may be measured over time using a amperemeter (in case of a constant voltage provided by the voltage source) or the voltage between the second electrode 42 and the third electrode 43 may be measured over time using a voltmeter (in case of a constant current provided by the voltage source, the method of measurement being termed cyclic voltammetry). The resulting time trace of the current or voltage may be used to analyze properties of the biomolecules, particularly nucleic acids, of the biological sample S. As an alternative, the properties of the biomolecules may also be analyzed by coulometry or by an impedance measurement.

[0185] Alternatively or additionally, the biomolecules in the biological sample (or amplified from the biological sample) may be analyzed by a marker molecule, which particularly may be disposed in the fibrous material F of the third layer 30 in dry or lyophilized form prior to entry of the biological sample S. In case of nucleic acids, the marker molecule may for instance be a hybridization probe such as a molecular beacon, which is capable of specifically binding to a target DNA or RNA sequence. When proteins are to be detected, the marker molecule may be e.g. an antibody, which specifically binds to an epitope in the protein of interest. The marker molecule may be coupled (e.g. covalently bound) to an optically detectable molecule, such as a dye or a fluorophore, which changes color or emits luminescence or fluorescence light upon specific binding of the marker to the biomolecule of interest. Alternatively, the marker may also comprise one or more substance(s) which undergoe(s) a chemical reaction resulting in a change of color or emission of fluorescent or luminescent light. In case of fluorescence, the device may be illuminated with light of wavelengths overlapping with the absorbance spectrum of the fluorophore, e.g. by an external light source.

[0186] To detect the light emitted by the marker, an external optical detector 80 may be used. An example of this is depicted in FIG. 7, where the optical detector 80 is a camera having a lens 81. In this case, the device 1 may be at least partially transparent to maximize the amount of detected light. To this end, for example, the cover layers 50 and/or cover 60 may be formed from a transparent material.

[0187] The device 1 and particularly the applied marker M may be configured, such that the emitted light may be detected by an off-the-shelf camera of a mobile device, such as a smart phone or a tablet computer. This improves usability of the device 1, particularly for applications outside of a laboratory, such as on-site analysis of environmental samples.

[0188] As an alternative to optical detection, marker molecule may also be detectable by its redox properties.

[0189] Of course, the described analysis of biomolecules in the biological sample S may be performed with and without previous electrophoresis.

[0190] FIG. 8 shows a further embodiment of a device 1 according to the invention. The device 1 differs from the device shown in FIG. 1, in that the device comprises a lid 61 instead of the cover 60, wherein the first electrode 41 (grid electrode) is arranged in or on the lid 61. The lid 61 may be attached to the upper cover layer 50 by a hinge 62. In particular, the first electrode 41 is printed onto the lid 61 (i.e. by applying electrically conductive ink). When the lid 61 is closed, the first electrode 41 is placed directly on the first area 2a of the first layer 10, such that electrophoresis of the biological sample S may be initiated by applying a potential difference between the first electrode 41 and the second electrode 42.

[0191] FIGS. 9 and 10 show embodiments of the device 1, where the heating elements 70 are formed as resistance heaters, more specifically as heating electrodes. The heating electrodes may be arranged on, particularly printed on (i.e. by applying an electrically conductive ink), the lower cover layer 50 (FIG. 9) or alternatively a further fourth layer 100 placed between the third layer 30 and the lower cover layer 50 (FIG. 10).

[0192] The heating electrodes comprise a grid 73 composed of parallel rods 74 surrounded by a rectangular frame 75 and contact tabs 71 connecting the grid 73 to an edge of the respective layer, where they can be connected at ends 72 to the poles of a voltage source to generate a current through the heating electrodes and generate heat due to the electrical resistance of the heating electrodes.

[0193] Such integrated heating electrodes obviate the need for an external heating device.

[0194] FIG. 11 shows a further device 1 according to the invention having a first layer 10, a second layer 20 and a third layer 30 of a fibrous material F as well as cover layers 50 and a cover 60. The device 1 is particularly identical to the embodiment shown in FIG. 1, except for the following differences: the first layer 10 comprises instead of a grid electrode a first electrode 41 shaped as an elongated tab positioned (in the view of FIG. 11) on the left edge of the first area 2a and the third layer 30 comprises only the second electrode 42 which is positioned on the right edge of the third area 2c. Furthermore, the second area 2b of the second layer 20 is off-set to the right in respect of the first area 2a of the first layer 10, and the third area 2c of the third layer 30 is offset to the right in respect of the second area 2b of the second layer 20. Thereby, the resulting sample path SP or channel formed by the fibrous material F of the first layer 10, second layer 20 and third layer 30 along the first axis A1 extends diagonally in respect of the vertical axis between the first electrode 41 and the second electrode 42. In particular, to move and electrophoretically separate negatively charged molecules in the biological sample, the first electrode 41 may be connected to a negative pole 92 of a voltage source and the second electrode 42 may be connected to a positive pole 91 of a voltage source. This results in a separation of the biological sample, whereby bands of molecules in the biological sample are aligned next to each other in a projection along the plane parallel to the second area 2b when the bands are detected optically.

TABLE-US-00001 List of reference numerals  1 Device for analyzing biological samples  2 Substrate  2a First area  2b Second area  2c Third area 10 First layer 11 Opening 20 Second layer 21 Opening 30 Third layer 41 First electrode  41a Contact tab  41b Grid stucture  41c Rod  41d Frame 42 Second electrode  42a Contact tab 43 Third electrode  43a Contact tab 50 Cover layer 60 Cover 61 Lid 62 Hinge 70 Heating element  70a Heating of the PCR reagents  70b Heating of the polymer gel 71 Contact tab 72 End 73 Grid 74 Rod 75 Frame 80 Optical detector 81 Lens 91 Positive pole 92 Negative pole 100  Fourth layer A1 First axis A2 Second axis B Barrier structure F Fibrous material H Polymer gel L Lysis agent P PCR reagents S Biological sample SP Sample path