JPWO2018065102A5 - - Google Patents

Description

The present invention relates to an analysis system according to the preamble of claim 1 and a method according to the preamble of claim 20.

Preferably, the invention is particularly preferably from humans or animals, particularly preferably for analysis and diagnosis of the presence of diseases and / or pathogens and / or for determining blood cell counts, antibodies, hormones, or steroids and the like. It discusses analyzing and inspecting samples. Therefore, the present invention is particularly in the field of bioanalytical methods. Food samples, environmental samples, or other samples can also optionally be tested, especially with respect to environmental analysis methods or food safety and / or to detect other substances.

In particular, the present invention can be used to determine, identify, or detect at least one analyte (target analyte ) of a nucleic acid product, such as a sample, preferably a particular nucleic acid sequence. In particular, the sample can be tested to determine at least one analyte qualitatively or quantitatively, for example so that disease and / or pathogens can be detected.

The present invention specifically discusses what is known as a point of care system, i.e., for samples at sampling sites and / or independently or away from a central laboratory. Discuss how to perform the inspection.

US 5,096,669 discloses a point of care system for testing biological samples, especially blood samples. The system includes disposable cartridges and analytical devices. With the sample received, the cartridge is inserted into the analytical device to perform the inspection. The cartridge includes a microfluidic system and a sensor device that includes electrodes, which are calibrated with a calibrator and then used to inspect the sample.

In addition, WO 2006/125767 A1 and US 2016/0047832 A1 are points for integrated and automated DNA or protein analysis, including disposable cartridges and analytical devices for fully automated processing and evaluation molecular diagnostic analysis using disposable cartridges. Disposable of the care system. The cartridge is designed to accept samples, especially blood, and in particular, cell disruption so that the bound PCR amplification product or nucleic acid sequence can be detected as a target analyte in what is known as a redox cycle treatment. It enables PCR and detection of PCR amplification products bound to capture molecules to which label enzymes have been added. The temperature in the PCR chamber can be manipulated by the associated Pelche element.

US 2009/0227476 A1 discloses a bioassay device capable of performing PCR in a PCR thermodynamic cycle chamber and subsequently detecting nucleic acids in a microarray. The PCR thermodynamic cycle chamber can be heated or cooled separately.

US 5,096,669 WO 2006/125767 A1 US 2016/0047832 A1 US 2009/0227476 A1

The problems addressed by the present invention enable or facilitate efficient, reliable, rapid, and / or as accurate as possible testing of samples, especially improved analytical systems for testing biological samples and To provide an improved method.

The above problems are solved by the analysis system according to claim 1 or by the method according to claim 20. A favorable form of development is the subject of the dependent claims.

In particular, the proposed analytical system for inspecting biological samples is to form a receiving cavity for receiving the sample and / or to amplify the analyte of the sample and / or to form an amplification product from the analyte of the sample. It preferably comprises at least one reaction cavity of.

Preferably, the analytical system is separate from the receiving and / or reaction cavities, especially for electrochemical detection of the analyte and / or amplification product preferably bound to the capture molecule of the sensor device. And / or includes a sensor device that is isolated from the receiving cavity and / or the reaction cavity.

According to a first aspect of the invention, the analytical system comprises an intermediate temperature control cavity for actively temperature controlling the amplified analyte and / or amplification product, particularly for heating, the intermediate temperature control cavity. Arranged between the receiving cavity and / or the reaction cavity on one side and the sensor device on the other side, and / or the sensor device is fluidly connected to the receiving cavity and / or the reaction cavity by an intermediate temperature control cavity. To.

Advantageously, intermediate temperature control cavities are used to avoid or reduce their unwanted hybridization before the analytes and / or amplification products are fed to the sensor device, and / or, in particular, the correspondence. Analysts and / or amplification products bound to each other such that the number of analytes and / or amplification products that are not attached to or available for hybridization to the capture molecule is maximized or at least increased. It is possible to separate or denature things. In this way, the yield of amplification product bound to the capture molecule and thus the efficiency of the test is increased.

In particular, an intermediate temperature control cavity is preferably used to allow the analyte and / or amplification product to be fed to the sensor device in a modified state and / or in a pre-temperature controlled state. It is possible to denature and / or temperature control the amplified analytes and / or amplification products within them before they are fed to the sensor device. Thus, the denaturation step of the analyte and / or amplification product in or on the sensor device can be omitted and / or the required temperature control of the amplification product in or on the sensor device can be reduced. As a result, the efficiency of inspection increases.

According to another aspect of the invention, which can also be performed independently, the analytical system, in particular in addition to the intermediate temperature control cavity, in particular, captures molecules, analytes , and / or amplifications from the sensor temperature controller through the sensor apparatus. Temperature control of trapped molecules and analysts and / or amplification products in the sensor device, especially for direct temperature control and / or so that heat is transferred to the product and vice versa. Includes a sensor temperature controller for.

Particularly preferably, the different temperatures of the trapped molecule and the analyte and / or the amplified product in or on the sensor device preferably correspond to the (different) analyte and / or the amplified product at different hybridization temperatures. Can be set or reacted using a sensor temperature controller so that it can be coupled to. Advantageously, the efficiency and / or specificity of testing and / or detection is therefore increased.

Preferably, the sensor device comprises a support, in particular a chip as a support, a plurality of electrodes arranged on the support, and / or a sensor compartment, and the sensor temperature control device is particularly heat-sensitive. The support and / or sensor compartment is preferably designed to control the temperature particularly directly, and / or the sensor temperature controller is such that it travels from the device through the support to the sensor compartment and vice versa. In the operating state, the sensor compartment is temperature-controlled from the outside and / or in a planar manner and / or centered on or relative to the support or its back so that heat is particularly transferred through the support. There is.

Achieve different temperature or temperature curves or temperature profiles within and / or within the sensor compartment using the sensor temperature controller and / or capture the amplification product against the trapped molecule (optimally). It is possible to adapt the temperature control of the support and / or sensor compartment to hybridize.

The analytical system of the present proposal is particularly portable, mobile, and / or a point of care system, and / or can be used specifically at a sampling site and / or away from a central laboratory.

The analytical system preferably comprises an analytical device and / or at least one cartridge for inspecting the sample.

The term "analytical device" is particularly mobile and / or can be used in the field and / or the sample or its components preferably in a cartridge and / or chemically and biologically with the cartridge. , And / or are preferably understood to mean instruments designed to be physically inspected and / or analyzed. In particular, the analytical device controls the inspection of the sample in the cartridge.

Particularly preferably, the analytical device is designed to accept or connect to a cartridge.

The term "cartridge" preferably accepts, stores, and physically, chemically, and / or organisms a sample to allow detection, identification, or determination of at least one analyte of the sample. It is preferably understood to mean a structural device or unit designed to be scientifically processed and / or prepared and / or measured.

Cartridges within the meaning of the present invention preferably include a fluid system having a plurality of channels, cavities, and / or valves for controlling flow through the channels and / or cavities.

In particular, within the meaning of the present invention, the cartridge is designed to be at least substantially flat, flat, and / or card-like, particularly as a (micro) fluidic card, and / or preferably closed. Designed as a body or container capable of, and / or a cartridge can be inserted and / or inserted into the analytical device of the present invention when it contains a sample.

The proposed method, especially for examining biological samples, particularly preferably to denature the analyte and / or the amplification product, and / or the undesired hybridization of the analyte and / or the amplification product to each other. Sample analytes and / or amplification products that are preferably amplified using PCR to avoid or reduce are hybridized between the reaction cavity and the sensor device and / or after amplification / replication and to capture molecules. Provided is active temperature control immediately prior to hybridization, especially heating.

Preferably, different analytes and / or amplification products or groups thereof are first produced, especially simultaneously and / or in parallel, using amplification reactions, especially PCR, in preferably different PCR chambers and / or reaction cavities. It is then sequentially attached to the capture molecule at different hybridization temperatures.

In particular, it provides that the first, second, and arbitrary third groups of amplification products are produced in different reaction cavities, but the analytes are in a common PCR chamber using amplification reactions, especially PCR. It can also be provided that and / or being amplified in the reaction cavity and / or that the amplification product is produced in a common reaction cavity.

Preferably, multiple amplification reactions, especially PCR, are performed simultaneously, in parallel or independently of each other during the test.

Preferably, different amplification reactions, especially PCR with different primers, are provided or carried out.

Within the meaning of the present invention, the amplification reaction is a particularly molecular-biological reaction in which the analyte is amplified / replicated and / or the amplification product, in particular the nucleic acid product of the analyte, is produced. Particularly preferably, PCR is an amplification reaction within the meaning of the present invention.

"PCR" is an abbreviation for the polymerase chain reaction and is a molecular-biological method that uses it to make a given analysis , especially a portion of the sample RNA or DNA, particularly an amplification product or a nucleic acid product. It is then amplified in several cycles, preferably using a polymerase or enzyme for testing and / or detection. If RNA is intended to be tested and / or amplified, cDNA is produced starting with RNA, especially using reverse transcriptase, before PCR is performed. The cDNA will be used as a template for the next PCR.

Preferably, during PCR, the sample is first denatured by the addition of heat to separate strands of DNA or cDNA. Preferably, the primer or nucleotide is then deposited on a single isolated strand of DNA or cDNA, the desired DNA or cDNA sequence is replicated with polymerase, and / or the lost strand is with polymerase. Will be replaced. This process is preferably repeated in multiple cycles until the desired amount of DNA or cDNA sequence is available.

For PCR, it is preferred to use marker primers, i.e., primers that produce (additionally) markers or labels, especially biotin-amplified analytes . This allows or facilitates detection. Preferably, the primers used are biotinylated and / or specifically contain or form covalent biotin as a label.

Analyzes and / or amplification products are preferably temperature controlled in advance and / or before (again) temperature control within the sensor device, especially any potentially double-stranded analyte and /. Or, preferably in an intermediate temperature control cavity and / or after exiting the reaction cavity and / or immediately before being fed to the sensor device and / or just before being fed to the sensor device to separate the amplification product into single chains. It is suggested that it be heated preferably (pre-) using an intermediate temperature controller.

Preferably, the analyte and / or amplification product is then temperature controlled in succession and / or again, especially after temperature control, especially in the intermediate temperature control cavity, and / or preferably corresponds to the amplification product. In order to hybridize to the trapped molecule, it is brought to the corresponding hybridization temperature, especially in and on the sensor device and / or using the sensor temperature controller.

Preferably, the sensor compartment, or capture molecule, analyte , and / or amplification product, is actively temperature controlled, especially by transferring heat through the support of the sensor device, especially the chip. / Or bring to the corresponding hybridization temperature.

The support is preferably contacted both electrically and thermally on the back surface and / or connected and / or coupled to the analytical device. This allows for a particularly compact design.

The hybridization temperature is preferably such that the (amplified) analyte, in particular a portion of RNA or DNA, and / or the amplification product is attached to the corresponding capture molecule and / or hybridized to the corresponding capture molecule. (Average) temperature.

The optimum hybridization temperature is preferably the temperature at which the number of amplification products bound to the corresponding capture molecule is maximized and / or the number of amplification products bound to each other is minimized.

Preferably, the (optimal) hybridization temperature varies for different analytes and / or amplification products.

The group of different analytes and / or amplification products preferably comprises, at least substantially, only the analytes and / or amplification products having similar (optimal) hybridization temperatures. Thus, this results in a temperature range of average and / or optimal hybridization temperature or (optimal) hybridization temperature of the group. At this temperature or in this temperature range of the group, both are abbreviated as "group temperature", but the total number of analytes and / or amplification products in this group bound to the capture molecule is maximum (maximum). there is a possibility). The temperature range is preferably less than 8 ° C, especially less than 5 ° C.

Preferably, different groups with different group temperatures are formed. Group temperatures are preferably greater than at least 2 ° C, especially 3 ° C, or are spaced apart.

In particular, the group temperature of the first group is higher than the group temperature of the second group.

Preferably, the (optimal) hybridization temperature is the GC content of the DNA or cDNA, the length of the DNA or cDNA, the melting or melting temperature of the DNA or cDNA sequence, and / or the solvent, ambient medium, and / or buffer. Varies according to adjustment or salt concentration.

The melting point or melting temperature is preferably the temperature at which the DNA or cDNA denatures from that temperature and / or the strands of double-stranded DNA or cDNA separate from each other. The melting point or melting temperature preferably depends on the GC content of the DNA or cDNA, the length of the DNA or cDNA, and / or the adjustment or salt concentration of the solvent, ambient medium, and / or buffer. Preferably, the melting or melting temperature is at least 85 ° C or 90 ° C, particularly preferably 92 ° C or 94 ° C, and / or maximum 99 ° C or 98 ° C, particularly preferably maximum 97 ° C or 96 ° C.

Preferably, the hybridization temperature is preferably at least 2 ° C. or 5 ° C., particularly preferably 8 ° C. or 10 ° C., particularly 15 ° C. or 20 ° C. or much lower than the melting point or melting temperature.

The capture molecule of the sensor device is a particularly oligonucleotide probe that is preferably immobilized by spacers, especially C6 spacers, on the sensor, sensor array, and / or electrodes. The formation of structures that interfere with hybridization, such as hairpin structures, can be avoided by the preferred binding of trapped molecules by spacers.

Analysts and / or amplification products combined at different hybridization temperatures are preferably detected by a single or common detection process.

Particularly preferably, the sensor device is used only once in the process for detecting the analyte and / or the amplification product, and the electrochemical determination is preferably for all of the coupled amplification products. On the other hand, it is done at the same time. This allows for very quick and efficient inspection.

In particular, different analytes and / or amplification products of different analytes are capable of measuring and / or determining or detecting a large number of different amplification products at the same time, especially in a single or common detection process. Particularly preferably, it is possible to bind to the trapped molecule, which is preferably immobilized on or in the sensor device, in order by the hybridization temperature very efficiently.

In the context of the present invention, it is therefore possible to inspect analytes and / or amplified products that are produced and / or amplified in parallel with high specificity at the same time in a single detection process and have different hybridization temperatures. ..

Preferably, the bound analyte and / or amplification product is detected by feeding the detector molecule to the sensor device.

Within the meaning of the present invention, the term "detector molecule" is specific to the marker or label of the primer used to amplify the analyte and / or the analyte or the amplification product produced with it. It is preferable to understand that it means a molecule that binds to and enables this detection.

In particular, the detector molecule can be enzyme-conjugated and / or immunoconjugated, including a reporter enzyme that specifically binds to markers or labels, especially biotin, and for converting substrates. In the context of the present invention, the detector molecule is a streptavidin with high affinity for biotin and / or an alkaline phosphatase capable of converting a non-reactive phosphate monoester into an electrochemically active molecule and a phosphate. It is preferable to be based.

Preferably, a detection system is used in which the label is based on biotin and the detector molecule is based on streptavidin / alkaline phosphatase. However, other detector molecules can also be used.

The aspects and features mentioned above of the present invention, as well as the claims and the aspects and features of the present invention revealed by the following description, can be practiced independently of each other in principle, but also in any combination or order. It can also be carried out at.

Other aspects, advantages, features, and properties of the invention will become apparent from the following description of preferred embodiments with reference to the claims and drawings.

FIG. 6 is a schematic cross-sectional view through the analytical system of the present proposal or through the analytical device in which the cartridge of the present proposal is accepted. It is a schematic diagram of a cartridge. It is a schematic front view of the sensor device of this proposal of an analysis system and / or a cartridge. It is an enlarged detailed view from FIG. 3 which shows the sensor field of a sensor device. It is a schematic rear view of a sensor device. It is the schematic sectional drawing of the sensor device. FIG. 5 is a schematic curve or profile diagram with respect to the temperature of the sample and / or amplification product as a function of cartridge position.

In these figures, which are only schematic and not at exact scale, the same reference symbols are used for the same or similar parts and components, and the corresponding or equivalent properties and advantages repeat these. It is achieved even if it is not explained.

FIG. 1 is a very schematic view of the analysis system 1 and analysis device 200 of the present proposal for inspecting a biological sample P, particularly preferably with or in a device or cartridge 100.

FIG. 2 is a schematic diagram of a preferred embodiment of the device or cartridge 100 of the present proposal for inspecting sample P. The device or cartridge 100 specifically forms a handheld unit, which is simply referred to below as the cartridge.

The term "sample" is preferably understood to mean a sample material to be tested, especially taken from a human or animal. In particular, within the meaning of the present invention, the sample is preferably a fluid or component thereof, such as saliva, blood, urine, or another liquid from a human or animal. To the extent of the meaning of the present invention, the sample can be pretreated or prepared as needed, or can be obtained directly from, for example, humans or animals. Food samples, environmental samples, or other substances, especially with respect to environmental analysis methods, food safety, and / or to detect other substances, preferably natural substances, but also biological or chemical weapons agents or poisons, etc. Samples can also be inspected arbitrarily.

Preferably, the analytical system 1 or analytical device 200 specifically controls and / or evaluates the inspection of the sample P in or on the cartridge 100, or collects, processes, and / or stores measurements from the inspection. Used for.

By using the proposed analysis system 1 or analysis device 200, or by using the cartridge 100, and / or by using the method of the present proposal for inspecting sample P, preferably analyte A of sample P. A nucleic acid sequence, such as a particular nucleic acid sequence, or particularly preferably a plurality of analytes A of sample P, can be determined, identified, or detected. In particular, these analytes are detected and / or measured not only qualitatively, but also preferably quantitatively.

Thus, in particular, sample P qualitatively or quantifies at least one Analyte A so that it can, for example, detect disease and / or pathogens, or determine other values that are important, for example, for diagnosis. Can be inspected to determine the target.

Particularly preferably, molecular biological testing is possible by using the analysis system 1 and / or the analysis device 200 and / or by using the cartridge 100.

Particularly preferably, molecular and / or PCR assays for detecting DNA and / or RNA, ie nucleic acid products and / or sequences, are possible and / or performed.

Preferably, the individual components of sample P or sample P or analyte A can be amplified as needed, especially using PCR, and tested in analysis system 1, analysis device 200, and / or cartridge 100. Can be identified and / or detected. Therefore, preferably, the amplification product V of the analysis product A or the analysis product A is produced.

Analyte A and / or amplification product V of sample P, especially nucleic acid products amplified by using PCR, are at least 20 or 50, particularly preferably 80 or 100, and / or up to 300 or 280. Particularly preferably it has a length of 250 or 220 nucleotides. However, it can also be provided that shorter or longer amplification product V is produced specifically by using PCR.

In the following, further details will first be given for the preferred configuration of the cartridge 100, and the features of the cartridge 100 will preferably be those of the analysis system 1, even if there is no further explicit description of any of them. It is expressed directly.

The cartridge 100 is preferably at least substantially flat, flat, and / or plate-shaped, and / or card-shaped.

The cartridge 100 preferably comprises at least a substantially flat, flat, plate-like, and / or card-like body 101, which body 101 is manufactured and / or injection molded from a particularly plastic material, particularly preferably polypropylene. To.

As shown by the broken line in FIG. 2, the cartridge 100 includes a main body 101 and / or at least a part thereof, and / or a valve for covering a cavity and / or a channel formed particularly on the front surface 100A. It preferably comprises at least one film or cover 102 for forming.

The analysis system 1 or the cartridge 100 or its body 101 preferably, together with the cover 102, preferably forms and / or includes a fluidic system 103, which is hereinafter referred to as a fluid system 103.

The cartridge 100 and / or its fluid system 103 is oriented, preferably at least substantially vertically, in the working position and / or during inspection, especially within the analytical device 200, as schematically shown in FIG. Thus, in particular, the spread of the principal plane or surface of the cartridge 100 extends at least substantially vertically in the working position.

The cartridge 100 and / or fluid system 103 includes a plurality of cavities, particularly at least one receiving cavity 104, at least one weighing cavity 105, at least one intermediate cavity 106, at least one mixing cavity 107, as shown in FIG. It preferably comprises at least one storage cavity 108, at least one reaction cavity 109, at least one intermediate temperature control cavity 110 and / or at least one recovery cavity 111.

The cartridge 100 and / or fluid system 103 more preferably comprises at least one pump device 112 and / or at least one sensor device 113.

Part, most, or all of the cavity is preferably formed by chambers and / or channels or other recesses within the cartridge 100 and / or body 101, and is particularly preferably covered or closed by a film or cover 102. The cover. But other structural solutions are possible.

In the illustrated example, the cartridge 100 or fluid system 103 has two weighing cavities 105A and 105B, a plurality of intermediate cavities 106A to 106G, a plurality of storage cavities 108A to 108E, and / or, as can be seen in FIG. It preferably comprises a plurality of reaction cavities 109 that can be loaded independently of each other, in particular a first reaction cavity 109A, a second reaction cavity 109B, and an optional third reaction cavity 109C.

The one / plurality of reaction cavities 109 are used, in particular, to carry out amplification reactions, especially PCR, or some preferably different amplification reactions, especially PCR. It is preferred that several preferably different PCRs, i.e., PCRs with different primer combinations or primer pairs be performed in parallel and / or independently and / or in different reaction cavities 109.

The amplification product V and / or other portion of sample P formed in one or more reaction cavities 109 can be guided or fed to the connected sensor device 113, especially using the pump device 112.

The sensor device 113 specifically detects the Analyte A of the Analyte A or the Sample P, in this case the amplification product V of the Analyte A, particularly preferably qualitatively and / or quantitatively. Used for. However, other values may be collected and / or determined in lieu of or in addition to this.

In particular, the pump device 112 includes or forms a tubular or beaded ridge, particularly preferably on the back surface of the cartridge, with a film or cover 102, as schematically shown in FIG.

The cartridge 100, body 101 and / or fluid system 103 preferably include a plurality of channels 114 and / or valves 115, as shown in FIG.

By using channels 114 and / or valves 115, cavities 104 to 111, pumping devices 112 and / or sensor devices 113, as needed, particularly as these are controlled by analysis system 1 or analysis device 200. / Or can optionally or selectively connect and / or separate from each other temporarily and / or permanently.

The cavities 104 to 111 are preferably fluid-communicated with each of the plurality of channels 114. Particularly preferably, each cavity is linked by at least two related channels 114 to allow the fluid to fill, flow through, and / or drain from each cavity as needed. Or connected.

The fluid transfer or fluid system 103 is preferably not based on or solely based on capillary forces, in particular gravity and / or pumping forces and / or pumping forces generated by the pump or pumping device 112. / Or is basically based on the effects of compressive and / or suction forces. In this case, fluid flow or fluid transfer and metering is done by opening and closing the valve 115 accordingly and / or in particular by operating the pump or pumping device 112 appropriately using the pump device 202 of the analytical device 200. Be controlled.

Preferably, each of the cavities 104-110 has an inlet at the top and an outlet at the bottom in the working position. Therefore, if necessary, only the liquid from each cavity can be taken out through the outlet.

In particular, liquid-containing cavities, particularly preferably storage cavities 108, mixing cavities 107 and / or receiving cavities 104, potentially cause gas or air bubbles to rise upward in the operating position when these cavities are filled with liquid. It can be formed, thereby sizing the liquid so that it collects above the outlet without air bubbles. However, in this case, other solutions are possible.

Preferably, at least one valve 115 is assigned to each cavity and the pump device 112 and / or the sensor device 113 is located upstream of each inlet and / or downstream of each outlet.

Preferably, the fluid can selectively open an arrangement of cavities 104 to 111 or cavities 104 to 111 through which the fluid flows, eg, sequentially or continuously, and / or by activating the assigned valve 115. Can selectively flow through and / or these cavities can be fluid connected to the fluid system 103 and / or other cavities.

In particular, the valve 115 is formed by the body 101 and the film or cover 102 and / or by another method, such as by an additional layer or recess.

Particularly preferably, the liquid or liquid reagent F in the storage cavity 108 and / or the fluid system 103 is particularly preferably sealed from the open receiving cavity 104 in a stable storage manner, preferably initially or in the storage state. One or more valves 115A are provided.

Preferably, the initial closure valve 115A is located upstream and downstream of each storage cavity 108. These valves are preferably opened particularly automatically only when the cartridge 100 is actually in use and / or during insertion of the cartridge 100 into the analytical device 200.

In this case, the supernatant of sample P, such as serum, can be optionally discarded or removed, especially when an optional intermediate connection 104D is provided in addition to the inlet 104B and outlet 104C. In addition, a plurality of valves 115A, particularly three valves, are preferably assigned to the receiving cavity 104. Based on the application, in addition to the valve 115A on the inlet 104B, preferably only the valve 115A at either the outlet 104C or the intermediate connection 104D is further opened.

The valve 115A assigned to the receiving cavity 104 particularly fluidly and / or the fluid system 103 and / or the cartridge 100 until the sample P is inserted and until the receiving cavity 104 or the connection 104A of the receiving cavity 104 is closed. Alternatively, seal it in an airtight manner.

One or two or more that are not closed in a stable storage manner and / or are initially open and / or can be closed by actuation as an alternative to or in addition to valve 115A (in initial closure). Valve 115B is preferably provided. These valves are used specifically to control fluid flow during inspection.

The cartridge 100 is preferably designed as a microfluidic card and / or the fluid system 103 is preferably designed as a microfluidic card. In the present invention, the term "microfluidic" refers to individual cavities, parts of cavities, or all of cavities 104 to 111 and / or channels 114, each having a volume of 5 ml separately or cumulatively. Alternatively, it is preferably understood to mean less than 2 ml, particularly preferably less than 1 ml or 800 μl, particularly less than 600 μl or 300 μl, and particularly more preferably less than 200 μl or 100 μl.

Particularly preferably, a sample P having a maximum volume of 5 ml, 2 ml, or 1 ml can be introduced into the cartridge 100 and / or the fluid system 103, especially the receiving cavity 104.

As shown in the schematic diagram shown in FIG. 2, a reagent preferably introduced or supplied in the form of a liquid or liquid reagent F in liquid form and / or in the form of dry reagent S prior to inspection for testing sample P. And liquid is needed.

In addition, for inspection, detection and / or other purposes, for example, in the form of a solvent and / or substrate SU, especially for wash buffers, drying reagents S, to form detection molecules and / or redox systems. Other liquids F in are also preferably required and are particularly supplied within the cartridge 100, i.e. also before use, especially before supply. Some of the following issues make no distinction between liquid reagents and other liquids, so the respective explanations can be applied to each other accordingly.

Analytical system 1 or cartridge 100 contains all reagents and liquids required to perform one or more amplification reactions or PCRs and / or tests, and thus particularly preferably optionally. It is only necessary to accept the preprocessed sample P.

The cartridge 100 and / or the fluid system 103 directs the sample P or its components directly to the sensor device 113 by passing through the reaction cavity 109 and bypassing the optional intermediate temperature control cavity 110 as needed. Specifically, when the bypass 114A is open to the sensor device 113 to feed the liquid or liquid reagent F2-F5 from the storage cavity 108B-108E so that it can be fed, the valve 115B of the bypass 114A It preferably comprises a bypass 114A that can be optionally used, especially when open, so that it can be transported or pumped in the opposite direction to the reagent A and / or the amplification product V.

The cartridge 100 or fluid system 103 or channel 114 preferably comprises a sensor portion 116 or other device for detecting liquid level and / or fluid flow.

The various components described in FIG. 2, such as the channel 114, the valve 115, in particular the valve 115A in the initially closed state, the valve 115B in the initially open state, and the sensor portion 116 are in some cases for clarity reasons. Note that the same symbol is used for each of these components in FIG. 2, although only labeled with.

The recovery cavity 111 is preferably used to receive excess or used reagents, liquids, and sample volumes. In particular, it is preferably provided with appropriately large dimensions and / or only inputs or inlets so that the liquid is not removed in the working position or pumped out again.

The receiving cavity 104 preferably includes a connecting portion 104A for introducing the sample P. After the sample P is introduced into the receiving cavity 104, the cavity and / or the connection 104A is closed.

The cartridge 100 can be inserted and / or accepted into the analytical device 200 of the present proposal to inspect the sample P, as shown in FIG. Instead, sample P can be supplied later.

FIG. 1 shows an analysis system 1 in a ready-to-use state for the stage of performing an inspection on the sample P received in the cartridge 100. Thus, in this state, the cartridge 100 is connected to, accepted by, and / or inserted into the analytical device 200.

In the following, some features and aspects of the analytical device 200 will first be described in more detail. The features and aspects of the device are themselves preferably also the features and aspects of the analysis system 1 of the present proposal, even in the absence of any further explicit description.

The analysis system 1 or analysis device 200 preferably includes a mount or receptacle 201 for mounting and / or accepting the cartridge 100.

Preferably, the cartridge 100 is fluidly, particularly hydraulically separated or isolated from the analytical device 200. In particular, the cartridge 100 forms a particularly closed fluid and / or hydraulic system 103 that is preferably independent of the sample P, reagents, and other liquids.

Preferably, the analytical device 200 is designed to operate the pump device 112 and / or the valve 115 specifically to detect thermal effects and / or measurement data using the sensor device 113 and / or the sensor portion 116. Will be done.

The analysis system 1 or analysis device 200 preferably includes, in particular, a pump drive 202 designed to mechanically activate the pump device 112.

Preferably, the head of the pump drive 202 can be rotated to push in the axial direction while rotating the preferably beaded raised portion of the pump device 112. Particularly preferably, the pump drive 202 and the pumping device 112 together form, in particular, a hose pump or peristaltic pump for the fluid system 103 and / or the cartridge 100, and / or a pump of the type metering pump.

Particularly preferably, the pump is configured as described in DE 10 2011 015 184 B4. However, other structural solutions are possible.

Preferably, the capacity and / or amount of discharge of the pump can be controlled and / or the feed direction of the pump and / or pump drive 202 can be switched. Thus, preferably, the fluid can be pumped in the forward or reverse direction as needed.

The analysis system 1 or analysis device 200 preferably includes a connecting device 203 for particularly electrically and / or thermally connecting the cartridge 100 and / or the sensor device 113.

As shown in FIG. 1, the connecting device 203 preferably includes a plurality of electrical contact elements 203A, and the cartridge 100, particularly the sensor device 113, is preferably electrically connected to the analytical device 200 by these contact elements 203A. Electrical connection is possible.

The analysis system 1 or analysis device 200 is for controlling the temperature of one or more temperature control devices 204, particularly the cartridge 100, and / or has a thermal effect on the cartridge 100, especially for heating and / or cooling. It preferably comprises a heating element or a Pelche element.

Individual temperature control devices 204, some of these devices, or all of these devices are preferably placed in contact with the cartridge 100, body 101, cover 102, sensor device 113, and / or individual cavities. Can and / or can be thermally coupled to them and / or can be incorporated therein and / or specifically actuated or electrically controlled by the analytical device 200. In the illustrated example, temperature control devices 204A, 204B, and / or 204C are provided in particular.

Preferably, the temperature control device 204A, which will be referred to below as the reaction temperature control device 204A, can carry out one or more amplification reactions and / or PCR in one or more reaction cavities 109, in particular inside them. Assigned as.

These reaction cavities 109 are preferably temperature controlled simultaneously and / or uniformly, particularly using one common reaction temperature control device 204A or two reaction temperature control devices 204A.

Particularly more preferably, the temperature of the one / plurality of reaction cavities 109 may be controlled using two reaction temperature control devices 204A preferably arranged from two different surfaces and / or on opposite surfaces. can.

Alternatively, the temperature of each reaction cavity 109 can be controlled independently and / or individually.

A temperature control device 204B, hereinafter referred to as an intermediate temperature control device 204B, is preferably assigned to the intermediate temperature control cavity 110 and / or preheats the intermediate temperature control cavity 110 or the fluid therein, particularly the amplification product V, preferably. Designed to control temperature up to temperature TV.

The intermediate temperature control cavity 110 and / or the intermediate temperature control device 204B particularly preferably feeds the fluids supplied to the sensor device 113, particularly the analyte A and / or the amplification product V, just before these fluids are supplied. It is preferably placed upstream or (directly) in front of the sensor device 113 so that the temperature can be controlled or preheated in the desired manner.

Particularly preferably, the intermediate temperature control cavity 110 and / or the temperature control device 204B denatures the sample P or the analyte A and / or the generated amplification product V and / or the double-stranded analyte A or amplification. It is designed or intended to split the product V into single chains and / or to prevent premature binding and / or hybridization of the amplified product V, especially due to the addition of heat.

The intermediate temperature control cavity 110 is preferably designed as a channel that extends and / or particularly winds or bends and / or has a flat cross section. Significantly, a sufficiently long retention time of the fluid and / or a sufficiently large thermal bond with the fluid in the intermediate temperature control cavity 110 is desired, for example, without changing the flow rate or while the fluid flows through the cavity. Obtained to achieve temperature control. However, other solutions, in particular where the flow of fluid in the intermediate temperature control cavity 110 is stopped, are also possible here.

Preferably, the length of the intermediate temperature control cavity 110 is at least 10 mm or 15 mm, particularly preferably at least 20 mm or 25 mm, particularly 30 mm or 40 mm, and / or maximum 80 mm or 75 mm, particularly preferably maximum 70 mm or 65 mm, in particular. The maximum is 60 mm.

Preferably, the intermediate temperature control cavity 110 is at least 10 μl or 20 μl, particularly preferably at least 25 μl or 20 μl, and / or at most 500 μl or 400 μl, and particularly preferably at most 350 μl or 300 μl.

The intermediate temperature control cavity 110 includes an inlet 110A and an outlet 110B, a valve 115, particularly preferably an initially open valve 115A assigned to (each) the inlet 110A and / or the outlet 110B as shown in FIG. Therefore, the flow of fluid through the intermediate temperature control cavity 110 can be controlled. For example, to temperature control the fluid flowing through the intermediate temperature control cavity 110 while flowing, and / or to stop and continue to pass the fluid in the intermediate temperature control cavity 110 for temperature control purposes only. The intermediate temperature control cavity 110 can be initially filled with a temperature controlled fluid and the input side and / or output side valve 115A can be closed.

Preferably, the intermediate temperature control cavity 110 is arranged (fluidically) between one reaction cavity / plurality of reaction cavities 109 and the sensor device 113, and / or (all) reaction cavities 109 are intermediate temperature controlled. The cavity 110 is preferably fluidly connected or connectable exclusively to the sensor device 113.

Preferably, the intermediate temperature control cavity 110 is located closer to the sensor device 113 than the one reaction cavity / plurality of reaction cavities 109. In particular, the distance or flow path between the intermediate temperature cavity 110, in particular the outlet 110B and the sensor device 113, is between the intermediate temperature control cavity 110, in particular the inlet 110A and one reaction cavity / plurality of reaction cavities 109. Shorter than distance or flow path.

The intermediate temperature control cavity 110 is preferably designed to heat the fluid, particularly the amplification product V, preferably to actively control the temperature to the melting point or melting temperature as described in detail below. To.

The intermediate temperature control device 204B assigned to the intermediate temperature control cavity 110 is preferably designed to (actively) control the temperature of the intermediate temperature control cavity 110, especially to heat it.

Preferably, the intermediate temperature controller 204B comprises or is formed by a heating element, particularly a heating resistor or a Pelche element.

The intermediate temperature control device 204B is preferably flat and / or has an elongated and / or rectangular contact surface that allows heat transfer between the intermediate temperature control device 204B and the intermediate temperature control cavity 110.

Preferably, the intermediate temperature control device 204B is external to the cartridge 100, the body 101, and / or the cover 102 in the region of the intermediate temperature control cavity 110 or on the intermediate temperature control cavity 110, preferably over the entire surface. Positioning from, especially can be pressed.

In particular, the analytical device 200 includes an intermediate temperature control device 204B. However, other structural solutions are also possible in which the intermediate temperature control device 204B is located within the cartridge 100 or integrated into the cartridge 100, especially the intermediate temperature control cavity 110.

Preferably, the analysis system 1, the analysis device 200 and / or the cartridge 100, and / or 1 or each temperature control device 204 specifically allows the temperature to be controlled and / or regulated in a temperature detector and / or Includes temperature sensor (not shown).

One or more temperature sensors can be assigned, for example, to the sensor portion 116 and / or individual channel portions or cavities, i.e., thermally coupled to them.

Particularly preferably, the temperature sensor is assigned to each temperature control device 204A, 204B, and / or 204C, for example, to measure the temperature of each temperature control device 204 and / or its contact surface.

The temperature control device 204C, hereinafter referred to as the sensor temperature control device 204C, is particularly assigned to the sensor device 113 and / or a fluid located in or on the sensor device, in particular the analyte A and / or the amplification product V or It is designed to control the temperature of the reagent or the like in a desired manner, preferably to the hybridization temperature TH.

The sensor temperature controller 204C preferably comprises or is formed by a heating element, particularly a heating resistor or a Pelche element.

The sensor temperature control device 204C is preferably flat and / or preferably rectangular and / or heat transfer between the sensor temperature control device 204C and the sensor device 113 corresponding to the dimensions of the sensor device 113. Has a contact surface that allows.

Preferably, the analytical device 200 includes a sensor temperature controller 204C. However, other structural solutions in which the sensor temperature control device 204C is incorporated within the cartridge 100, particularly the sensor device 113, are also possible.

Particularly preferably, the connecting device 203 includes the sensor temperature control device 204C, and / or the connecting device 203 can be connected to the cartridge 100, particularly the sensor device 113, together with the sensor temperature control device 204C, and can be particularly pressed.

Particularly more preferably, the connecting device 203 and the sensor temperature control device 204C (together) to electrically bond the analytical device 200 to the cartridge 100, particularly to the sensor device 113 or its support 113D, and to thermally couple with it. Can be moved towards and / or relative to the cartridge 100, in particular the sensor device 113, and / or can be placed in contact with the cartridge.

Preferably, the sensor temperature control device 204C is centrally located on the connecting device 203 or its support and / or between the contact elements 203A.

In particular, the contact element 203A is connected such that the connecting device 203 is thermally connected to or connectable to the sensor device 113 at the center and preferably outside the edge region or within the edge region. It is located in the edge region of device 203 or its support or around sensor temperature control device 204C. However, in this case, other solutions are possible.

The analysis system 1 or analysis device 200 preferably includes one or more actuators 205 for activating the valve 115. Particularly preferably, various actuators 205A and 205B assigned to these valves are provided to activate each of the various valves 115A and 115B, respectively.

The analysis system 1 or analysis device 200 preferably includes one or more sensors 206. In particular, the sensor 206A is designed or intended to detect liquid level and / or fluid flow within the fluid system 103. Particularly preferably, the sensor 206A determines the liquid level and / or its presence, velocity, mass flow rate / volumetric flow rate, temperature, and / or other values of the fluid in the channel and / or cavity, especially in the plane and / or plane of the fluid system 103. Or designed to measure or detect, for example, optically and / or capacitively, within each assigned sensor portion 116 formed by the extended channel portion.

Particularly preferably, each of the sensor portions 116 allows the fluid to vertically and / or from bottom to top at the operating position of the cartridge 100 to allow or facilitate reliable detection of the liquid. Directed to flow through and / or incorporated within the fluid system 103, and / or fluid thus flows in contact with or through the sensor portion 116.

Alternatively or additionally, the analytical device 200 adjusts the ambient temperature, internal temperature, atmospheric humidity, position, and / or alignment, for example, by a GPS sensor and / or the orientation and / or orientation of the analytical device 200 and / or cartridge 100. / Or preferably includes a (other or additional) sensor 206B for detection using tilt.

The analysis system 1 or the analysis device 200 collects, in particular, measurements and / or other data or values from and / or from the inspection results, in particular to control the sequence of inspections. It preferably comprises a controller 207 that includes an internal clock or time reference for evaluation and / or output or supply.

The control device 207 preferably prefers the pump drive 202, the temperature control device 204, and / or the actuator 205 in consideration of or depending on the desired inspection and / or the measurements from the sensor devices 113 and / or the sensor 206. Controls or adjusts.

In general, the cartridge 100, the fluid system 103, and / or the fluid feed does not preferably operate on the basis of capillary forces, but at least basically or primarily due to the effects of gravity and / or the pump or pumping device 112. Note that it works under the effect.

In the operating position, the liquid from each cavity is removed in each case through the outlet at the bottom, especially preferably withdrawn, especially through the inlet at the top where gas or air flows into and / or pumps. It can be sent. In particular, the associated vacuum in the cavity can be thus prevented or at least minimized when feeding the liquid.

The flow of fluid is controlled, in particular, by invoking the pump or pumping device 112 accordingly and further invoking the valve 115.

Particularly preferably, the pump drive 202 includes a stepper motor or drive that is calibrated in a separate manner so that proper activation can at least result in the desired weighing in principle.

In addition to or in place of this, the sensor 206A is specifically assigned a sensor to provide the desired fluid sequence and desired metering by controlling the pump or pumping device 112 accordingly and further invoking the valve 115 accordingly. It is preferred that the liquid level or fluid flow be detected in cooperation with the portion 116.

Optionally, the analysis system 1 or the analysis device 200 includes an input device 208 such as a keyboard or touch screen, and / or a display device 209 such as a screen.

The analysis system 1 or analysis device 200 is at least one for controlling, communicating, and / or outputting measurement data or test results, and / or linking to other devices such as printers or external power supplies. Two interfaces 210 are preferably included. This interface can be, in particular, a wired or wireless interface 210.

The analysis system 1 or analysis device 200 preferably includes a power source 211, preferably a built-in and / or externally connected or externally connectable battery or storage battery.

Preferably, the built-in storage battery is provided as a power source 211 and is (re) charged and / or replaceable by an external charging device (not shown) through the connection 211A.

The analysis system 1 or analysis device 200 preferably includes a housing 212 in which all components and / or some or all of the devices are preferably incorporated. Particularly preferably, the cartridge 100 can be inserted or slipped into the housing 212 through an opening 213, such as a slot that can be closed, and / or can be accepted by the analytical device 200.

The analysis system 1 or analysis device 200 is preferably portable or mobile. Particularly preferably, the analytical device 200 is lighter than 25 kg or 20 kg, particularly preferably lighter than 15 kg or 10 kg, and particularly lighter than 9 kg or 6 kg.

The details of the preferable configuration of the sensor device 113 are given below with reference to FIGS. 3 to 6.

The sensor device 113 preferably enables electrochemical measurements and / or redox cycles.

In particular, the sensor device 113 identifies and detects an Analyte A (equal or different) bound (equal to or different) to the capture molecule M or a product derived from it, in particular the amplified product V of the Analyte A or various Analysts A. , And / or designed to determine.

The sensor device 113 preferably includes a sensor array 113A including a plurality of sensor regions or sensor fields 113B as shown in a schematic diagram in FIG. 3, which shows the measurement side of the sensor device 113 and / or the sensor array 113A. FIG. 4 is an enlarged detailed view from FIG. FIG. 5 shows the connection side, and FIG. 6 is a schematic cross-sectional view passing through the sensor device 113.

Preferably, the sensor device 113 or sensor array 113A is more than 10 or 20, particularly preferably more than 50 or 80, particularly more than 100 or 120, and / or 1000 or Includes less than 800 sensor fields 113B.

Preferably, the sensor device 113 or the sensor array 113A includes a plurality of electrodes 113C. It is preferable that at least two electrodes 113C are arranged in each sensor region or sensor field 113B. In particular, in each case at least two electrodes 113C form the sensor field 113B.

The electrode 113C is preferably made of a metal, particularly a noble metal such as platinum or gold, and / or the electrode is particularly coated with a thiol.

As can be seen from the enlarged detail view of the sensor field 113B described in FIG. 4, preferably the electrodes 113C are finger-shaped and / or engage with each other. However, other structural solutions or sequences are also possible.

The sensor device 113 preferably comprises a support 113D, particularly a chip, and the electrode 113C is preferably located on and / or incorporated within the support 113D.

The measurement side includes the electrode 113C and / or is the side facing the fluid, sample P, amplification product V, and / or sensor compartment, and / or the analyte A and / or the amplification product V is coupled. This is the side of the sensor device 113 and / or the support 113D that includes the capture molecule M (shown in FIG. 6).

The connection surface of the sensor device 113 and / or the support 113D is preferably the opposite side of the measurement surface and / or the side facing away from the fluid, sample P, and / or amplification product V.

Particularly preferably, each of the measurement side and the connection side of the sensor device 113 and / or the support 113D forms one flat side surface of the support 113D, which is particularly flat and / or plate-like.

The sensor device 113, particularly the support 113D, preferably includes a plurality, in this case eight electrical contacts or contact surfaces 113E, which are located on the connecting side and / or connected as shown in FIG. It is preferable to form the side.

Preferably, the sensor device 113 can be brought into contact with the connecting side and / or using the contact 113E and / or can be electrically connected to the analytical device 200. In particular, the electrical connection can be established by electrically connecting the contact 113E between the cartridge 100, in particular the sensor device 113 and the analytical device 200, in particular the control device 207, to the contact element 203A.

Preferably, the contacts 113E are arranged laterally around the edge region and / or the electrodes 113C and / or the sensor array 113A in plan or projection, and / or the contacts 113E are the edges of the sensor device 113. As far as possible from the region, especially the support 113D electrically, preferably laterally as described above by using the connecting device 203 or the contact element 203A, and / or in the center or the support 113D. It extends so that it can be electrically contacted around the sensor temperature controller 204C, which can be preferably positioned in the upper center.

Preferably, the sensor fields 113B are separated from each other as shown in the schematic of FIG. In particular, the sensor device 113 includes a barrier or partition between each of the sensor fields 113B, preferably formed by a particularly hydrophobic layer 113F having a corresponding recess in the sensor field 113B. However, other structural solutions are possible.

The cartridge 100 and / or the sensor device 113 includes or forms a sensor compartment 113G. In particular, the sensor compartment 113G is formed between the sensor array 113A, the sensor device 113, and / or the support 113D, or between the measurement surface on one side and the sensor cover 113H on the other side.

The sensor device 113 preferably defines the sensor compartment 113G by using the measurement surface of the sensor device and / or the sensor array 113A. Therefore, the electrode 113C is in the sensor compartment 113G.

Preferably, the cartridge 100 and / or the sensor device 113 includes a sensor cover 113H, particularly a sensor compartment 113G defined or delimited by the flat side sensor cover 113H.

Particularly preferably, the sensor cover 113H can be lowered onto the partition and / or layer 113F for actual measurement.

The fluid system 103, so that the sensor device 113 or the sensor compartment 113G can accept the (preprocessed) sample P, analyte A, or amplification product V to the measurement side of the sensor device 113 or sensor array 113A. In particular, it is fluidly linked to one reaction cavity / plurality of reaction cavities 109 by a connection 113J.

Thus, the sensor compartment 113G can be loaded with a fluid and / or this fluid can flow through the sensor compartment.

The sensor device 113 preferably comprises a plurality of, in particular, different capture molecules M, wherein the different capture molecules M are preferably located and / or immobilized and / or different sensors in or on the different sensor fields 113B. It is preferably assigned to field 113B.

Particularly preferably, the capture molecule M is provided to electrode 113C through bond B, especially a thiol bond, in this case to bind and / or detect or identify particularly suitable analyte A and / or amplification product V. ..

Different capture molecules M1 to M3 have different sensor fields 113B and / or different sensor fields 113B and / or to specifically bind different analytes A and / or amplification products V, in FIG. 6, amplification products V1 to V3 within sensor field 113B. It is preferably provided for different electrode pairs and / or electrodes 113C.

Particularly preferably, the sensor device 113 or the sensor array 113A allows the coupled amplification product V to be determined qualitatively or quantitatively within each sensor field 113B.

Preferably, the sensor device 113 comprises a capture molecule M having a different hybridization temperature TH, preferably to bind the amplification product V to the corresponding capture molecule M at a different hybridization temperature TH.

To achieve hybridization of different hybridization temperatures TH, the temperature of the sensor device 113, in particular the electrode 113C, the support 113D, the sensor compartment 113G, and / or the cover 113H, is at least indirectly, preferably the analytical device 200. In particular, it can be controlled or set using the sensor temperature control devices 204B and / or 204C as described above.

Preferably, the sensor temperature controller 204C, in this case, is brought into contact with the connecting surface so that the particularly desired or required hybridization temperature TH reaches the sensor compartment 113G and / or the measurement plane in the fluid. It is used to control the temperature of 113G.

Preferably, in the operating state, the sensor temperature controller 204C is planar and / or centered and / or opposite to the sensor array 113A and / or at least partially resting on one or more contacts 113E. Places on support 113D. This allows, in particular, rapid and efficient temperature control of the sensor compartment 113G and / or amplification product V.

The sensor device 113, particularly the support 113D, is at least one, preferably a plurality of electronic circuits or integrated, specifically designed to detect the current or voltage preferably generated in the sensor field 113B according to the redox cycle principle. The circuit is preferably included.

Particularly preferably, the measurement signals from the different sensor fields 113B are collected or measured separately by the sensor device 113 and / or their circuits.

Particularly preferably, the sensor device 113 and / or the integrated circuit directly converts the measurement signal into a digital signal or data that can be read, in particular by the analytical device 200.

Particularly preferably, the sensor device 113 and / or the support 113D is configured as described in EP 1 636 599 B1.

In the following, a preferred sequence of inspection or analysis using the proposed analysis system 1 and / or the analysis device 200 and / or the proposed cartridge 100 and / or following the method of the present proposal will be described in more detail below.

The analysis system 1, the cartridge 100 and / or the analysis device 200 are preferably designed to carry out the method of the present proposal.

In the proposed method of inspecting sample P, at least one analyte A of sample P is preferably amplified or replicated, especially using PCR. The amplified analyte A and / or the amplified product V produced thereby binds and / or hybridizes to the corresponding capture molecule M. The bound amplification product V is detected, especially using electrical measurements.

The method can be used to detect diseases and / or pathogens, especially in the field of medicine, especially veterinary medicine.

Within the context of the methods according to the invention, a sample P having at least one Analytical A based on a fluid or liquid from the human or animal body, particularly blood, saliva, or urine, is for detecting disease and / or pathogens. Usually first introduced into the receiving cavity 104 through the connection 104A, the sample P can be pretreated.

With the sample P accepted, the receiving cavity 104 and / or its connection 104A is fluid-closed in a liquid-tight and / or airtight manner.

Preferably, the cartridge 100, along with the sample P, is then linked or connected to the analytical device 200 and, in particular, inserted or slipped into the analytical device 200.

Method sequences, in particular fluid flow and feed and mixing, etc., are initiated by the analytical device 200 or control device 207, in particular with the pump drive 202 or pump device 112 and / or the actuator 205 or valve 115 actuated accordingly. Is controlled by.

Preferably, the sample P or a portion or supernatant of the sample P is removed from the receiving cavity 104 through the outlet 104C and / or through the intermediate connection 104D and supplied to the mixing cavity 107 in a weighing manner.

Preferably, the sample P in the cartridge 100 is weighed specifically in or with the first metering cavity 105A and / or the second metering cavity 105B before being introduced into the mixing cavity 107. In this case, particularly upstream and / or downstream sensor portions 116 are used in combination with the assigned sensor 206 to allow desirable weighing. However, other solutions are possible.

Sample P is prepared for yet another analysis in the mixing cavity 107 and / or preferably fed into the liquid reagent F1 from the first storage cavity 108A and / or into the mixing cavity 107. It is mixed with one or more drying reagents S1, S2, and / or S3.

Liquid reagents and / or dry reagents can be introduced into the mixing cavity 107 before and / or after sample P. In the illustrated example, the drying reagents S1 to S3 are preferably pre-introduced into the mixing cavity 107 and optionally dissolved by sample P and / or liquid reagent F1.

The liquid reagent F1 can be a reagent in particular, especially a PCR master mixture for amplification reactions or PCR. Preferably, the PCR master mixture is nuclease-free water, enzymes for performing PCR, especially at least one DNA polymerase, nucleoside triphosphate (NTP), especially deoxynucleotide (dNTP), salts, especially magnesium chloride, and / Or contains a reaction buffer.

The drying reagents S1, S2, and / or S3 can also be reagents required to carry out an amplification reaction or PCR and are in a dry state, especially in a lyophilized form. Preferably, the drying reagents S1, S2, and / or S3 are particularly selected from lyophilized enzymes, preferably DNA polymerase, NTP, dNTP, and / or salts, preferably magnesium chloride.

Dissolution or mixing within the mixing cavity 107 is generated or facilitated, especially by introducing and / or blowing gas or air from below. This introduction is carried out, in particular, by pumping gas or air appropriately into the circuit using a pump or pumping device 112.

Subsequently, a desired volume of sample P mixed and / or pretreated in the mixing cavity 107 is added to one or more reaction cavities 109, particularly preferably different reagents or primers, in this case drying reagents S4 to S6. One (respectively) of any upstream intermediate cavities 106A to 106C added or dissolved is particularly preferably fed through to one or more reaction cavities 109.

Particularly preferably, the (premixed) sample P is preferably divided into several sample portions of equal size and / or preferably evenly and / or of equal size sample portions of intermediate cavities 106A to 106C. Distribute between and / or between reaction cavities 109.

Different reagents, in this case drying reagents S4 to S6, particularly preferably primers, particularly those required for one or more PCRs, especially in this case a group of different primers, react differently with intermediate cavities 106A to 106C and / or different. It is preferably added to sample P (premixed) within each of the cavities 109.

The primers in the different groups differ with respect to the hybridization temperature of the amplification product V generated by each primer. As a result, different group temperatures, especially in the group of Analyte A and / or Amplified Product V, are produced as mentioned at the beginning.

Particularly preferably, a marker primer is used in the sense already specified at the beginning.

In the illustrated embodiment, reagents or primers S4 to S6 are contained within intermediate cavities 106A to 106C. However, other solutions, in particular reagents or primers S4 to S6, may be contained within the reaction cavity 109.

According to a preferred embodiment, each of the intermediate cavities 106A through 106C comprises a primer for amplifying / replicating one analyte A, preferably two different analytes A, more preferably three different analytes A. However, 4 or 5 or more different analytes A can be amplified / replicated for each reaction cavity 109.

Particularly preferably, the reaction cavities 109 are sequentially filled with a specified volume of (pretreated) sample P or each sample portion through intermediate cavities 106A to 106C, each located upstream. For example, the first reaction cavity 109A is filled with a designated volume of pretreated sample P prior to the second reaction cavity 109B, and / or the second reaction cavity 109B is of the third reaction cavity 109C. Filled with this sample before.

In the reaction cavity 109, an amplification reaction or PCR is performed to replicate / amplify Analyte A. These reactions were performed using a particularly assigned and preferably common reaction temperature controller 204A and / or preferably simultaneously for all reaction cavities 109, i.e., particularly the same circulation and / or temperature (curve / profile). ) Is used.

One or more PCRs are basically performed based on a protocol or temperature profile known to those of skill in the art. In particular, the volume of the mixture or sample in the reaction cavity 109 is preferably periodically heated and cooled.

Preferably, a nucleic acid product is produced from Analyte A as Amplified Product V in one / a plurality of reaction cavities 109.

During pretreatment, reaction, and / or PCR or amplification, label L is produced directly (in each case) and / or attached to amplification product V. This is achieved in particular by using the corresponding preferably biotinylated primers. However, the label L can be produced separately or later optionally for the first time in the sensor compartment 113G and / or after hybridization and / or bound to the amplification product V.

Label L is specifically used to detect bound amplification product V. In particular, the label L can be detected or identified in the detection process, as described in more detail below.

According to the present invention, a large number of (different) analytes A can be replicated or amplified in parallel and multiple amplification reactions or PCRs can be performed in parallel using different primers S4 to S6 and / or primer pairs so that they can be analyzed later. And / or can be carried out independently of each other.

In particular, preferably using amplification reactions, especially PCR performed in parallel, the same or different analytes A1 are amplified in the first reaction cavity 109A and the same or different analytes A2 are in the second reaction cavity 109B. The same or different analyte A3 is amplified in the third reaction cavity 109C.

Particularly preferably, the analytes A1 to A3 are different from each other so that a particularly large amount of different analytes A can be amplified and / or inspected using the methods of the invention. Preferably, Analyte A of more than 2 or 4, particularly preferably more than 8 or 11, especially more than 14 or 17, can be inspected and / or amplified particularly simultaneously.

In particular, multiple groups of amplification product V of Analyte A are preferably formed and / or produced in parallel and / or independently of each other and / or in the reaction cavity 109. Thus, for example, a first group of amplification products V1 of analyte A1 is formed and / or produced in the first reaction cavity 109A, and a second group of amplification products V2 of analyte A2 is second. A third group of amplification products V3 of Analyte A3 is formed and / or formed in any third reaction cavity 109C.

Particularly preferably, a group of (amplified) analyte A and / or amplification product V having different group temperatures in the sense mentioned at the beginning is formed. Therefore, this group preferably has different (optimal) hybridization temperature THs and / or hybridization temperature ranges.

Preferably, different groups of analyte A and / or amplification product V, in particular nucleic acid products and / or sequences, are thus amplified and / or formed for testing, and different groups are amplified and / or The formed and / or particularly different reaction chambers 109A to 109C are fed, but instead are fed in different fashions.

After performing PCR and / or amplification, the corresponding fluid volume and / or amplification product V and / or group is specifically group-specific and / or separate intermediate cavities 106E, 106F, or 106G from reaction cavity 109, respectively. ) And / or through an optional (common) intermediate temperature control cavity 110 to the sensor device 113 and / or the sensor compartment 113G in sequence.

Intermediate cavities 106E-106G are yet another reagent for preparing amplification product V for hybridization, in this case drying reagents S9 and S10, respectively, eg, buffers for yet another preparation, especially SSC. May contain buffers and / or salts. Based on this, yet another adjustment of amplification product V can be performed, especially to improve the efficiency of subsequent hybridization (binding to capture molecule M). Particularly preferably, the pH of sample P is set or optimized within 106G from the intermediate cavity 106E and / or with the drying reagents S9 and S10.

Preferably, the group formed by the sample P or the analyte A and / or the amplification product V or the amplification product V is particularly just before being fed between the sensor device 113 and / or the reaction cavity 109 and the sensor device 113. In particular, the temperature is actively controlled using and / or in the intermediate temperature control cavity 110 and / or using the intermediate temperature control device 204B (especially in advance and / or in the sensor device 113). Before), preferably preheated.

Preferably, the analyte A or amplification product V of the group and / or individual reaction cavities 109 is actively temperature controlled (especially in advance and / or before temperature controlled by the sensor device 113) and / Alternatively, they are sequentially supplied to the intermediate temperature control cavity 110. These groups are particularly supplied to the sensor device 113, and / or the sensor compartment 113G is temperature controlled in sequence, especially prior to and / or before being temperature controlled by the sensor device 113.

FIG. 7 shows an exemplary schematic curve or profile of the temperature T of sample P as a function of position X in or above the cartridge 100.

Sample P is fed to the cartridge 100 and / or the receiving cavity 104 at or with an ambient temperature TU, such as about 20 ° C. The amplification reaction is subsequently carried out in the reaction cavity 109, preferably circulating heating and cooling the pretreated sample P (not shown in FIG. 7).

As mentioned above, a plurality of groups having particularly different analytes A and / or amplification product V and / or group temperature or hybridization temperature TH are preferably produced.

The group and / or amplification product V is preferably fed from the assigned intermediate cavity 106E to 106G and / or to the next intermediate temperature control cavity 110, preferably in sequence.

Preferably, the group and / or amplification product V is cooled at different rates and / or continuously in the reaction cavity 109, and / or the group and / or amplification product V is shown schematicly in FIG. As such, they exit the reaction cavity 109 in sequence and / or at different temperatures. However, preferably the group and / or the amplification product V is temperature controlled so that the group and / or the amplification product V exits the reaction cavity 109 at the same temperature and / or is constant within the reaction cavity 109 after completion of PCR. Other method transformations that are kept at temperature are also possible.

Preferably, the group and / or amplification product V is cooled on the way to the intermediate temperature control cavity 110. In this process, the group and / or amplification product V is shown in FIG. 7 between the reaction cavity 109 and the inlet 110A of the intermediate temperature control cavity 110 and / or by jumping the temperature curve or temperature profile of the reaction cavity 106. By absorbing the reagents S9 and S10 contained in these cavities as described above, it is possible to cool the intermediate cavities 106B to 106G particularly significantly and / or additionally.

Preferably, the group and / or amplification product V is at inlet 110A of intermediate temperature control cavity 110, as shown in FIG. 7 at inlet 110A, especially when they leave reaction cavities 109A and 109C at different temperatures. It has a different inlet temperature TE. However, the inlet temperature TE can also be substantially the same.

The inlet temperature TE of the inlet 110A is preferably at least substantially corresponding to the ambient temperature TU or up to 10 ° C. or 5 ° C. above the ambient temperature TU. However, the inlet temperature TE can be higher if necessary.

Preferably, the group and / or amplification product V is heated (in order) in the intermediate temperature control cavity 110 to the preheating temperature TV and / or the melting point or melting temperature, and the preheating temperature TV is the outlet of the intermediate temperature control cavity 110. It is preferable to reach at 110B (at the latest).

Preferably, the preheating temperature TV is higher than the hybridization temperature TH, especially at least as high as the melting point or melting temperature of each group and / or amplification product V. In particular, the group and / or amplification product V denatures the group and / or amplification product V as described above, especially just before being fed between the sensor device 113 and / or the reaction cavity 109 and the sensor device 113. Therefore, it is heated to the preheating temperature TV.

As shown in FIG. 7, all of the group and / or amplification product V are preferably heated to the same preheating temperature TV, eg at least 95 ° C.

However, other methods in which the group and / or amplification product V is temperature controlled (especially prior and / or before temperature controlled by the sensor device 113) and / or heated to a different preheating temperature TV (pre). It can also be transformed. In particular, the preheating temperature TV can be varied for each group and / or depending on the desired hybridization temperature TH and / or group temperature. In particular, the preheating temperature TV of the first group can be higher than the preheating temperature TV of the second and / or third group, and / or the preheating temperature TV can be lowered for each group.

The melting point or melting temperature and / or preheating temperature TV is preferably above the respective hybridization temperature TH, and / or in particular such that the bonds of analyte A and / or amplification product V are formed by intermediate dissolution. And / or at least 70 ° C. or 80 ° C. and / or at maximum 99 ° C. or 96 ° C. so that Analyte A and / or Amplified Product V is fed to sensor device 113 in a modified and / or dissolved state. be.

Optionally, the group of analytes A and / or amplification product V and / or amplification product V is temperature controlled (especially in advance and / or before temperature control by the sensor device 113), preferably these. Is (preliminarily) heated to the corresponding hybridization temperature TH, especially before being fed to the sensor device 113, so that it can bind directly to the corresponding capture molecule M after being fed to the sensor device 113.

In an alternative method variant, the group and / or amplification product V is actively temperature controlled, specifically heated (alone) in or on sensor device 113, and / or preferably sensor temperature control device 204C only. Is raised to the corresponding hybridization temperature TH. In particular, both denaturation of any hybridized amplification product V and (next) hybridization of the amplification product V and the corresponding capture molecule M can occur within or on the sensor device 113. .. In this case, therefore, the previous (intermediate) temperature control in front of the sensor device 113 can be omitted.

In a preferred method variant, however, sample P and / or group or analyte A and / or amplification product V is active, especially just before being fed to sensor device 113 and / or between reaction cavity 109 and sensor device 113. Temperature controlled (especially in advance and / or before temperature controlled in sensor device 113) and / or preferably raised to preheating temperature TV using intermediate temperature control device 204B to sensor device 113. After being fed and / or subsequently and / or again temperature controlled within the sensor device 113 (especially after temperature control in the intermediate temperature control cavity 110) and / or preferably the sensor temperature control device 204C. It is used to raise the corresponding hybridization temperature TH and / or group temperature. In this case, thus any hybridized amplification product V is denatured before and / or outside the sensor device 113 before being fed to the sensor device 113.

In particular, in a preferred method modification, the sample P and / or group and / or amplification product V rapidly exits the respective hybridization temperatures TH and / or multiple reaction cavities 109 at multiple stages or after exiting one reaction cavity / plurality of reaction cavities 109. / Or raised to group temperature, preferably the amplification product V is temperature controlled in the first stage, especially in the intermediate temperature control cavity 110 and / or in advance and / or in the sensor device 113. Temperature controlled above the hybridization temperature TH and / or up to the preheating temperature TV and / or modified at melting point or melting temperature, followed by and / or again temperature controlled in the second step, especially the corresponding It is heated and / or cooled down to the hybridization temperature TH and / or group temperature, particularly in the sensor device 113 and / or after temperature control in the intermediate temperature control cavity 110.

In particular, in this case the sensor temperature so as to avoid unwanted cooling below the respective hybridization temperature TH and / or group temperature of the preheated sample P and / or group in the intermediate temperature control cavity 110. In particular, the sensor device 113 is preheated using the control device 204C.

Particularly preferably, the sensor device 113 is at least substantially each up to the hybridization temperature TH of each Analyte A and / or amplification product V and / or up to their respective group temperatures, or slightly higher or lower temperatures. If preheated. Due to the relatively large heat capacity of the sensor device 113, preferably warm samples P and / or groups are fed to the sensor device 113 and / or its sensor compartment 113G to the desired and / or optimum temperature for hybridization ( Can be reached (rapidly).

The amplification product V, nucleic acid product, and / or group from the reaction cavity 109 are sequentially introduced into the sensor device 113 as detected or determined specifically within the sensor device 113.

Preferably, the first group and / or amplification product V1 from the first cavity 109A is fed to the sensor device 113 and / or the second group and / or amplification from the second reaction cavity 109B. The second group and / or amplification product V2 from the second reaction cavity 109B is bound to the corresponding capture molecule M1 prior to the product V2, in particular the third group from the third reaction cavity 109C. And / or is attached to the corresponding capture molecule M2 prior to the amplification product V3.

After the sample P and / or the amplification product V is fed to the sensor device 113, the amplification product V hybridizes to the capture molecule M.

In the context of the present invention, it has been demonstrated that it is particularly advantageous to hybridize the group of amplification product V and / or amplification product V at a hybridization temperature TH and / or group temperature specifically selected in each case. Has been done.

Particularly preferably, sample moieties and / or amplification product V from different PCRs and / or from different reaction cavities 109 are captured at different hybridization temperatures TH and / or lower hybridization temperatures TH and / or group temperatures. It binds to the molecule M in particular order.

Preferably, each of the analyte A and / or the amplification product V within the group has a similar, preferably at least substantially the same (optimal) hybridization temperature TH, to which they bind to the appropriate capture molecule M. However, the analyte A and / or the amplification product V within the group can also have a somewhat different (optimal) hybridization temperature TH, i.e., a range of hybridization temperatures as described at the outset. Thus, this results in a temperature range of the average and / or optimal hybridization temperature TH of the group or the (optimal) hybridization temperature of this group. This hybridization temperature or this temperature range of the group is sometimes shortly referred to as the "group temperature".

The group and / or amplification product V can hybridize at a descending or ascending, preferably descending group temperature and / or hybridization temperature TH. When the descending hybridization temperature TH is used, the already bound amplification product V can be kept from the capture molecule M by the next temperature increase.

Particularly preferably, the group temperature and / or the hybridization temperature TH1 of the first group, the amplification product V1 and / or the analysis product A1 is the group of the second group, the amplification product V2 and / or the analysis product A2. The temperature and / or the hybridization temperature is higher than TH2, which is higher than the third group, the amplification product V3, and / or the third group temperature and / or the hybridization temperature TH3 of the analyte A3.

It is understood that "hybridization temperature" specifically means the temperature at which, on average, the most analyte A and / or amplification product V in each group binds to the appropriate capture molecule M.

The group temperature and / or hybridization temperature TH of the different groups is preferably greater than 3 ° C., particularly greater than 4 ° C., preferably about 5 ° C. or greater.

Preferably, the group temperature and / or hybridization temperature TH is at least 40 ° C. or 45 ° C. and / or at most 75 ° C. or 70 ° C.

Preferably, the first group temperature and / or hybridization temperature TH1 of the first group and / or amplification product V1 is at least 55 ° C. or 58 ° C., particularly preferably at least 60 ° C. or 62 ° C., and / or maximum. 80 ° C. or 78 ° C., particularly preferably 75 ° C. or 72 ° C. at the maximum.

Preferably, the second group temperature and / or hybridization temperature TH2 of the second group and / or amplification product V2 is at least 40 ° C. or 45 ° C., particularly preferably at least 48 ° C. or 52 ° C., and / or maximum. 70 ° C. or 65 ° C., particularly preferably 60 ° C. or 58 ° C. at the maximum.

Preferably, the third group and / or the third group temperature and / or hybridization temperature TH3 of the amplification product V3 is at least 35 ° C. or 40 ° C., particularly preferably at least 42 ° C. or 45 ° C., and / or maximum. 65 ° C. or 62 ° C., particularly preferably 60 ° C. or 55 ° C. at the maximum.

At a first group temperature and / or hybridization temperature TH1, such as about 60 ° C., the first group binds particularly well to the corresponding or suitable capture molecule M1. At a second group temperature and / or hybridization temperature TH2, such as about 55 ° C., the second group binds particularly well to the corresponding or suitable capture molecule M2. At a third group temperature and / or hybridization temperature TH3, such as about 50 ° C., the third group binds particularly well to the corresponding or suitable capture molecule M3.

As shown in FIG. 7, the group and / or amplification product V is cooled from the intermediate temperature control cavity 110 in the middle of the sensor device 113. Preheating temperature TV and / or temperature predominates when fluid enters sensor device 113, and / or group temperature and / or optimal hybridization temperature, depending on temperature control in the prior and / or intermediate temperature control cavity 110. Depending on the TH, individual or all groups and / or amplification product V or sensor device 113 will need to be temperature controlled to different ranges using sensor temperature control device 204C.

For example, the first group and / or amplification product V1 from the first reaction cavity 109A is temperature controlled and in particular the second group and / or amplification product V2 and / or from the second reaction cavity 109B. Alternatively, it is heated or cooled to a higher degree than the third group and / or amplification product V3 from the third reaction cavity 109C.

In particular, the temperature control of the sensor device 113, in particular the support 113D, is applied to each group and / or different amplification product V to reach the respective group temperature and / or hybridization temperature TH.

In the illustrated example, the hybridization temperature TH1 of the first group is above the temperature of the first group entering the sensor device 113, which is preferably the first group from the first reaction cavity 109A. And / or the amplification product V1 must be heated, eg, 2 ° C. or 5 ° C., for greater hybridization.

However, the hybridization temperature TH can also correspond to the inlet temperature TE or the temperature of the input to the sensor device 113. In this case, each group and / or amplification product V is kept at a constant temperature in or on the sensor device 113 for hybridization. In particular, the group and / or amplification product V is the sensor device 113 at the corresponding hybridization temperature TH to the second group and / or amplification product V2 from the second reaction cavity 109B as shown in FIG. May be supplied in advance.

The inlet temperature TE or the temperature of the input to the sensor device 113 can be higher than the hybridization temperature TH of each group and / or amplification product V. In this case, each group and / or amplification product V has a temperature relative to the third group and / or amplification product V3 from the third reaction cavity 109C as shown in FIG. 7 at a particularly specified rate. Cooling or (slightly) temperature controlled in or on the sensor device 113 for hybridization so that it drops to the desired hybridization temperature TH.

According to the present invention, the hybridization temperature TH is changed stepwise, for example, in increments of several degrees Celsius and / or in increments of 5 ° C., and the hybridization temperature TH is group or at least one analyte A and / or amplified production. Both continuous and / or gradual changes, especially reductions, can be provided during hybridization of the substance V.

In another method modification, the temperature of each group and / or each group of amplification product V is controlled differently, and / or preferably different amplification product V of each group is hybridized differently. It can be provided that the temperature changes in or on the sensor device 113 for hybridization of the amplification product V in one of the groups so that it binds to the corresponding capture molecule M at temperature TH.

Sample P, group, analyte A and / or amplification product V hybridized and / or bound to capture molecule M, particularly preferably with the attached label L or in another scheme. The detection continues with.

Although particularly preferred detection variants, specifically electrochemical detection, will be described below in more detail, other types of detection, such as optical detection or capacitive detection, can be performed.

Following each binding / hybridization, preferably an optional wash treatment is performed and / or additional reagents or liquids are optionally fed from storage cavities 108B to 108E, in particular.

In particular, providing removal of sample residues and / or unbound amplification product V, reagents, and / or residues from PCR, and other substances that may interfere with the rest of the method sequence. can.

Washing or flushing can be carried out using, in particular, a fluid and / or reagent F3 preferably contained in the reservoir 108C, particularly a wash buffer, particularly preferably a sodium citrate buffer or an SSC buffer. Unbound analyte A and / or amplification product V, and material that may interfere with subsequent detection, are preferably removed from sensor device 113 by wash buffer and / or fed to recovery cavity 111. To.

Subsequent and / or subsequent cleaning treatments are followed by detection of amplification product V bound to capture molecule M according to a preferred modification of the method.

In order to detect the amplification product V bound to the capture molecule M, the reagent F4 and / or the detector molecule D, particularly alkaline phosphatase / streptavidin, is preferably supplied from the storage cavity 108D to the sensor device 113.

As shown in FIG. 6, the reagent F4 and / or the detector molecule D can bind to the bound amplification product V, particularly the label L of the bound amplification product V, particularly preferably the biotin marker.

In the context of detection, it can be provided that additional liquid reagents F3 and / or F5 are fed from storage cavities 108C and / or 108E to sensor device 113.

Optionally, following or after the reagent F4 and / or the detector molecule D binds to the amplification product V and / or the label L, especially the unbound reagent F4 and / or the detector molecule from the sensor device 113. A (additional) wash treatment and / or flush is performed, preferably with a fluid and / or reagent F3 and / or wash buffer to remove D.

Preferably, the reagents S7 and / or S8 and / or the substrate SU for detection are suitable for the substrate SU, particularly preferably from the reservoir 106B which is suitable for dissolving the reagents S7 and / or S8 and / or the substrate SU. The fluid or reagent F2 (particularly buffer) to be sampled is finally fed to the sensor device 113, especially from the storage cavity 106D, together with it. In particular, reagents S7 and / or S8 can form or contain substrate SU.

After adding the substrate SU, the cover 113H is preferably lowered to separate the sensor fields 113B from each other and / or to minimize substrate exchange during this time.

Preferably, p-aminophenyl phosphate (pAPP) is used as the substrate SU.

The substrate SU preferably reacts with the bound amplification product V and / or the detector molecule D and / or reacts with them and / or allows them to be electrochemically measured. ..

Preferably, the substrate SU is due to the alkaline phosphatase of the bound detector molecule D, particularly the bound detector molecule D, preferably such as p-aminophenol having particularly electrochemical and / or redox activity. It is preferably divided into a first substance SA and a second substance SP such as phosphate.

Preferably, the first or electrochemically active substance SA is detected by electrochemical measurements and / or redox cycles within the sensor device 113 or individual sensor fields 113B.

Particularly preferably, with the first substance SA, a redox reaction occurs particularly at the electrode 113C, and the first substance SA preferably emits or accepts electrons to or from the electrode 113C.

In particular, the presence and / or amount of the first substance SA in each sensor field 113B is detected by the associated redox reaction. That is, it is possible to qualitatively and particularly quantitatively determine whether or not the analyte A and / or the amplification product V is bound to the capture molecule M and how many are bound in each sensor field 113B. can. It provides information about which Analytes A were or was present in Sample P, and in particular information about the amount of these Analysts A.

In particular, due to the redox reaction with the first substance SA, a current signal or an electric signal is generated at the assigned electrode 113C, and this current signal or the electric signal is preferably detected by using the assigned electronic circuit. ..

Based on the current signal or electrical signal from the electrode 113C thus generated, whether or not hybridization with the capture molecule M has occurred and / or where it has occurred is determined.

The measurements are preferably made only once and / or for the entire sensor array 113A and / or for all of the sensor fields 113B, especially simultaneously or in parallel. In particular, the combined groups and / or amplification product V from all of the groups and / or reaction cavities 109 are detected, identified, or determined in a single or common detection process simultaneously or in parallel.

In other words, rapid measurements are possible, and regardless of this, high specificity for hybridization of the analyte A and / or the amplification product V with the capture molecule M is set in the manner targeted in each case. Amplification product V from individual reaction cavities 109 bound at different and / or specifically selected hybridization temperatures TH, together and / or as achieved based on the hybridization temperature TH. Detected in parallel.

However, in principle, it is also possible to measure a plurality of sample portions sequentially or separately in the sensor device 113 or in the plurality of sensor devices 113.

The inspection result or measurement result is particularly electrically transmitted to the analysis device 200 or its control device 207, preferably using the electrical connection device 203, and is prepared, analyzed, stored, in particular by the display device 209 and / or the interface 210. Displayed and / or output.

After the inspection has been performed, the cartridge 100 is cut from and / or released from and / or released from the analytical device 200 and is particularly discarded.

The individual aspects and features of the invention, as well as the individual method steps and / or method modifications, can be carried out independently of each other, but also in any desirable combination and / or order.

In particular, the invention also relates to any one of the following aspects that can be realized independently or in any combination and in combination with any of the aspects described above.

1. 1. The analysis system (1) provides a receiving cavity (104) for the sample (P) and / or a reaction cavity (109) for forming an amplification product (V) of the analyte (A) of the sample (P). It also includes a sensor device (113) for detecting the analyte (A) and / or the amplification product (V), the sensor device (113) including the receiving cavity (104) and / or the reaction cavity (109). An analytical system (1) for inspecting a biological sample (P), which is fluidly connected to the analytical system (1), the analytical product (A) and / or the amplified product (V). An intermediate temperature control cavity (110) is included for active temperature control, and the intermediate temperature control cavity (110) is a receiving cavity (104) on one side and / or a reaction cavity (109) and a sensor on the other side. An analytical system (113C) comprising a support (113D) and a plurality of electrodes (113C) placed between the device (113) and / or the sensor device (113). 1) is characterized by including a sensor temperature control device (204C) for directly controlling the temperature of the support (113D).

2. The analytical system (1) includes a plurality of reaction cavities (109) for producing the amplification products (V) in parallel and / or independently, and / or the sensor device (113) includes the amplification products (113). It comprises a capture molecule (M) for binding V) and is characterized in that the sensor device (113) is preferably fluidly connected to all reaction cavities (109) through the intermediate temperature control cavity (110). The analysis system according to the first aspect.

3. 3. An analytical system according to aspect 1 or 2, wherein the intermediate temperature control cavity (110) is elongated and / or preferably designed as a wavy channel.

4. The analysis system (1) includes an intermediate temperature control device (204B) for actively controlling the temperature of the intermediate temperature control cavity (110), preferably the intermediate temperature control device (204B) is a heating resistor or a Perche element. An analytical system according to any of the prior embodiments comprising or formed by the invention.

5. A support (113D) is placed between the electrodes (113C) and the sensor temperature control device (204C), and in the operating state, the sensor temperature control device (204C) is preferably flat on the support (113D). An analysis system according to any one of the preceding embodiments, wherein and / or centered and / or a sensor temperature controller (204C) comprises or is formed by a heating resistor or a Pelche element.

6. The support (113D) comprises or is formed by a chip, the chip is preferably electrically contactable, and / or a plurality of electrical contacts (113E), particularly laterally, in the edge region. And / or an analytical system according to any one of the preceding embodiments comprising around a sensor temperature controller (204C).

7. The analysis system (1) includes a connecting device (203) for electrically and / or thermally connecting the sensor device (113), particularly the support (113D), preferably the connecting device (203). Includes sensor temperature controller (204C) and / or multiple electrical contact elements (203A) and / or can move, particularly press against sensor device (113), especially supports (113D), or An analysis system according to any one of the preceding embodiments, characterized in that the reverse is also true.

8. The analysis system (1) includes a cartridge (100) for receiving the sample (P) and an analysis device (200) for receiving the cartridge (100), preferably the cartridge (100) is the receiving cavity (104). Includes one / plurality of reaction cavities (109), sensor device (113), and / or intermediate temperature control cavity (119) and / or analytical device (200) is sensor temperature control device (204C), intermediate. An analytical system according to any one of the preceding embodiments comprising a temperature control device (204B) and / or a connecting device (203).

9. The analyte (A) of the sample (P) is pretreated and / or the amplification product (V) is generated from the analyte (A) of the sample (P) in the reaction cavity (109) and pretreated. The analyzed article (A) and / or the amplified product (V) was bound to the capture molecule (M) on the support (113D) of the sensor device (113), and the bound analyte (A) and / or Alternatively, the amplification product (V) is a method of inspecting a biological sample (P) detected using the sensor device (113), and the amplification product (V) is the reaction cavity (109) and the sensor device. Actively temperature controlled between (113) and / or temperature controlled capture molecule (M) and / or analyte (A) and / or amplification product (V) and / or corresponding high A method characterized in that the support (113D) is directly temperature controlled so as to reach the hybridization temperature (TH).

10. After leaving the reaction cavity (109), the amplification product (V) is brought to the hybridization temperature (TH) in multiple steps and / or is actively temperature controlled in advance, preferably a sensor device (preferably a sensor device). Immediately prior to 113), it is preheated in the intermediate temperature control cavity (110), especially above the hybridization temperature (TH) and / or at least 70 ° C or 80 ° C and / or up to 99 ° C or 95 ° C. And / or subsequently according to embodiment 9, characterized in that the temperature is controlled in or on the sensor device (113) and / or the temperature is controlled to the corresponding hybridization temperature (TH), particularly heating and / or cooling. Method.

11. Different analytes (A) are preferably amplified in parallel and / or independently of each other and / or in multiple reaction cavities (109) and / or different from the first group of amplification products (V). The method according to aspect 9 or 10, characterized in that a second group of amplification products (V) is formed preferably in parallel and / or independently of each other and / or in different reaction cavities (109).

12. The amplification product (V) and / or the first and second groups are in turn fed to the sensor device (113) and / or in turn bound to the corresponding capture molecule (M) and / or The method according to aspect 11, characterized in that it is detected or determined by a single or common detection process.

13. The amplification product (V) and / or the first and second groups are actively temperature controlled and preferably heated as the fluid flows, especially to denature the amplification product (V). A method according to any one of aspects 9 to 12, characterized in that.

14. Any of aspects 9 to 13, characterized in that the amplification product (V) and / or the first and second groups are attached to the corresponding capture molecule (M) at different hybridization temperatures (TH). One way.

15. Aspect 9 characterized in that the analyte (A) is amplified using an amplification reaction, in particular PCR, and / or the nucleic acid product is produced from the analyte (A) as an amplification product (V). Method by any one of 14 to 14.

List of Reference Codes 1 Analysis System 100 Cartridge 100A Front 101 Body 102 Cover 103 Fluid System 104 Receiving Cavity 104A Connection 104B Inlet 104C Outlet 104D Intermediate Connection 105 Weighing Cavity 105A First Weighing Cavity 105B Second Weighing Cavity 106 (A) -G) Intermediate Cavity 107 Mixing Cavity 108 (A to E) Storage Cavity 109 Reaction Cavity 109A First Reaction Cavity 109B Second Reaction Cavity 109C Third Reaction Cavity 110 Intermediate Temperature Control Cavity 110A Inlet 110B Outlet 111 Connection Cavity 112 Pump device 113 Sensor device 113A Sensor array 113B Sensor field 113C Electrode 113D Support 113E Contact 113F Layer 114 Channel 114A Bypass 115 Valve 115A Initial closing valve 115B Initial opening valve 116 Sensor part 117 Sensor cover 118 Sensor compartment 119 Inlet 120 Outlet 200 Analytical device 201 Receptacle 202 Pump drive 203 Connecting device 203A Contact element 204 Temperature control device 204A Reaction temperature control device 204B Intermediate temperature control device 204C Sensor temperature control device 205 (Valve) Actuator 205A (Valve) actuator 205B 115B (Valve) actuator 206 Sensor 206A Fluid sensor 206B Other sensor 207 Controller 208 Input device 209 Display 210 Interface 211 Power supply 211A Connection 212 Housing 213 Opening A (1-3) Analytical object
B Bond D Detector molecule F (1-5) Liquid reagent L Label M (1-3) Capture molecule P Sample S (1-10) Drying reagent SU Substrate SA First substance SP Second substance T Temperature TE inlet Temperature TH (1-3) Hybridization temperature TU Ambient temperature TV Preheating temperature V (1-3) Amplification product X position

Claims (14)

This is a method for inspecting a biological sample (P).
The sample (P) divided into a plurality of sample portions is distributed among the plurality of reaction cavities (109).
A group of amplification products (V) are generated from the analyte (A) of sample (P) in each reaction cavity (109).
After performing the amplification, each of the groups of amplification products (V) is sequentially flushed from each of the reaction cavities (109) through the intermediate temperature control cavity (110) to the sensor device (113). ,
The amplification product (V) is bound to the capture molecule (M) on the support (113D) of the sensor device (113), and the bound amplification product (V) is the sensor device (113). In the method detected using
Each of the groups of amplification products (V) is preheated by the intermediate temperature control device (204B) in the intermediate temperature control cavity (110) between the respective reaction cavities (109) and the sensor device (113). The temperature is actively controlled up to the temperature,
The group of amplification products (V) each have a different inlet temperature at the inlet of the intermediate temperature control cavity (110) and are heated or heated to the same preheating temperature at the intermediate temperature control cavity (110).
The preheating temperature is varied for each group of amplification products (V) .
A method characterized by that. The method of claim 1, wherein the amplification product (V) is preheated in an intermediate temperature control cavity (110) immediately preceding the sensor device (113). The amplification product (V) is actively temperature controlled up to a temperature above the hybridization temperature (TH) and / or the amplification product (V) is.
The method of claim 2, wherein the temperature is controlled in or on the sensor device (113) and / or to the corresponding hybridization temperature (TH). A different analyte (A)
In parallel and / or independently of each other and / or in multiple reaction cavities (109)
The method according to any one of claims 1 to 3, wherein the method is amplified. The first group of amplification products (V) and the second group of different amplification products (V) are
In parallel and / or independently of each other and / or in different reaction cavities (109)
The method according to any one of claims 1 to 4, characterized in that it is formed. The first group and the second group are sequentially supplied to the sensor device (113) and / or are sequentially bound to the corresponding capture molecule (M), respectively. Item 5. The method according to Item 5. The method according to claim 5 or 6, wherein the first group and the second group are detected, identified, or determined by a single or common detection process. 1. The group of amplification products (V) is sequentially supplied to the sensor device (113) and / or is sequentially bound to the corresponding capture molecule (M), respectively. The method according to any one of claims 7. The method according to any one of claims 1 to 8, wherein the amplification product (V) is detected, identified, or determined by a single or common detection process. The amplification product (V) according to any one of claims 1 to 9, wherein the amplification product (V) is heated as the fluid flows through the amplification product (V ) in order to denature the amplification product (V). Method. The aspect according to any one of claims 1 to 10, wherein the amplification product (V) is bound to the corresponding capture molecule (M) at different hybridization temperatures (TH). the method of. The support (113D) is directly temperatureed to control the temperature of the capture molecule (M) and / or the amplification product (V) and / or to reach the corresponding hybridization temperature (TH). The method according to any one of claims 1 to 11, wherein the method is controlled. A claim that the analyte (A) is amplified using an amplification reaction and / or a nucleic acid product is produced from the analyte (A) as an amplification product (V). The method according to any one of claims 1 to 12. The method according to any one of claims 1 to 13, wherein the preheating temperature is higher than the hybridization temperature.