Cartridge and methods for removal or mixing of the first sample fraction from filter membrane

Abstract

The present invention relates to a cartridge for the uptake and analysis of a fluid sample, comprising an inlet which allows to take up the sample; a filter zone comprising a filter membrane which allows for separating different components within the sample and for adding reagents to the sample; a landing zone which allows for delaying the movement of the sample and for blending the sample; a transport zone which allows for transporting the sample from the landing zone to a reaction chamber, comprising one or more fluidic channels; one or more reaction chambers, which are designed for mixing of reagents with the sample; one or more reaction chambers, which are designed to capture analytes from the sample and other reactants; a reaction chamber zone, which is designed for washing, examination and verification of assay reactions; and one or more stop zones where fluidic flow within the cartridge is stopped.

Description

TECHNICAL FIELD

[0001] The present invention relates to a cartridge for the uptake and analysis of a fluid sample, comprising an inlet which allows to take up the sample; a filter zone comprising a filter membrane which allows for separating different components within the sample and for adding reagents to the sample; a landing zone which allows for delaying the movement of the sample and for blending the sample; a transport zone which allows for transporting the sample from the landing zone to a reaction chamber, comprising one or more fluidic channels; one or more reaction chambers, which are designed for mixing of reagents with the sample; one or more reaction chambers, which are designed to capture analytes from the sample and other reactants; a reaction chamber zone, which is designed for washing, examination and verification of assay reactions; and one or more stop zones where fluidic flow within the cartridge is stopped.BACKGROUND

[0002] Point Of Care Testing (POCT) can provide results to a physician more rapidly than routine central lab testing. Thus, substituting central lab testing with that performed at the POC is considered to increase speed and efficiency of diagnostic approaches. In particular, for patient safety, proper analytical validation and establishing high-sensitivity performance is critical. Handheld devices which integrate several functions such as sample preparation, analyte detection and signal processing are hence of paramount importance for medical practitioners, in particular in remote locations without central lab environments. However, the architecture of handheld devices may bring about certain errors, in particular spot-to-spot errors which lead to situations in which the first portion of a sample, e.g. blood, being analyzed is different from subsequent portions. There is hence a need for an improved device architecture which avoids the problem and increases analysis efficiency.SUMMARY

[0003] The present invention addresses this need and provides a cartridge for the uptake and analysis of a fluid sample, comprising: (i) an inlet which allows to take up the sample; (ii) a filter zone comprising a filter membrane which allows for separating different components within the sample and for adding reagents to the sample; (iii) a landing zone which allows for delaying the movement of the sample and for blending the sample; (iv) a transport zone which allows for transporting the sample from the landing zone to a reaction chamber, comprising one or more fluidic channels; (v) one or more reaction chambers, which are designed for mixing of reagents with the sample; (vi) one or more reaction chambers, which are designed to capture analytes from the sample and other reactants; (vii) a reaction chamber zone, which is designed for washing, examination and verification of assay reactions; and (viii) one or more stop zones where fluidic flow within the cartridge is stopped. It is particularly preferred that the landing zone of the cartridge comprises a structured baseplate with a fluidic channel that is designed to collect and transport filtered sample material, wherein said structured baseplate is arranged below the filter membrane. A correspondingly shaped landing zone is advantageously capable of decreasing spot to spot errors. In particular, the structure of the landing zone, specifically the baseplate leads to mixing of the sample material with reagent material and thereby drastically reduces spot-to-spot differences.

[0004] In a preferred embodiment of the present invention, said structured baseplate comprises a multitude of elevations.

[0005] In a particularly preferred embodiment, said elevations are formed as round or rectangular structures. In a further particularly preferred embodiment the elevations are pillar structures.

[0006] In another embodiment the structured baseplate comprises a multitude of contact points with the filter membrane, allowing for an efficient filtration into the landing zone.

[0007] In a further embodiment, the structured baseplate comprises a fluidic stop, venting and air system.

[0008] In a particularly preferred embodiment said fluidic stop, venting and air system comprises edges in the baseplate material, wherein said edges have a radius of < 0.02 mm.

[0009] In yet another embodiment, the filter membrane comprises pores of a diameter of 1.6 to 2.8 um.

[0010] In yet another preferred embodiment the landing zone comprises a surface coated with a hydrophilic material and / or wherein said elevations of the structured baseplate comprises a surface coated with hydrophilic material.

[0011] It is particularly preferred that the hydrophilic material is non-biofouling.

[0012] In a further preferred embodiment, the non-biofouling material additionally comprises a fluorescence dye.

[0013] In another embodiment hydrophilic material or non-biofouling material does not touch the edges in the baseplate material.

[0014] In a further aspect the present invention relates to a system comprising a handheld device capable of operating the cartridge and the cartridge as defined herein.

[0015] In a further aspect the present invention relates to a method of producing a cartridge as defined herein, comprising the steps: (a) preparing a structured baseplate as defined herein, preferably in a molding process; (b) preparing a filter membrane as defined herein by depositing one or more reagents on the filter membrane; and (c) depositing reagents in zones and chambers as defined herein.

[0016] In a further aspect the present invention relates to a method for uptaking and analyzing a fluid sample in a cartridge as defined herein, comprising at least the steps of: (a) entering a sample to the cartridge; (b) blocking the further penetration at a landing zone; (c) analyzing the sample in a reaction chamber zone; and (d) registering values measured during analysis for subsequent analysis or diagnostic steps.

[0017] In a particularly preferred embodiment of the cartridge, system or method of the present invention said fluid sample is a blood or blood plasma sample.

[0018] In yet another aspect the present invention relates to a use of the cartridge as defined herein or the system as defined herein for molecular diagnosis, biological sample analysis, preferably blood sample analysis, pharmacology, biological or chemical sample analysis, food analysis, environmental, veterinarian or forensic analysis.

[0019] It is to be understood that the features mentioned above and those yet to be explained below may be used not only in the respective combinations indicated, but also in other combinations or in isolation without departing from the scope of the invention.BRIEF DESCRIPTION OF THE DRAWINGS

[0020] FIG. 1 shows a schematic illustration of certain embodiments of components of a cartridge according to the invention. Certain components of the cartridge are shown in an explosion view on the left-hand side, including different layers such including a baseplate (1), a laminate layer (2) and a filter (3) . On the right hand-side a schematic illustration of bottom section (1) is provided in an enlarged view. The bottom section comprises a filter support (4) which is surrounded by a recess (5). Further depicted is a transport zone (6) and a reaction chamber zone (7) . Further indicated is a landing zone (8) covering the filter support (4), recess (5) and an initial section of transport zone (6). Enlarged views of the landing zone in Fig. 2 to 4 are indicated with II-IV. FIG. 2 to 4 show enlarged schematic illustrations of different landing zone versions (round, rectangular, oval) comprising a filter support (4) with an adjacent recess (area surrounding the filter support), elevations (10) and a fluidic channel that transports the liquid to reaction chambers, shown as (11). DETAILED DESCRIPTION OF EMBODIMENTS

[0021] Although the present invention will be described with respect to particular embodiments, this description is not to be construed in a limiting sense.

[0022] Before describing in detail exemplary embodiments of the present invention, definitions important for understanding the present invention are given.

[0023] As used in this specification and in the appended claims, the singular forms of "a" and "an" also include the respective plurals unless the context clearly dictates otherwise.

[0024] In the context of the present invention, the terms "about" and "approximately" denote an interval of accuracy that a person skilled in the art will understand to still ensure the technical effect of the feature in question. The term typically indicates a deviation from the indicated numerical value of ±20 %, preferably ±15 %, more preferably ±10 %, and even more preferably ±5 %.

[0025] It is to be understood that the term "comprising" is not limiting. For the purposes of the present invention the term "consisting of" or "essentially consisting of" is considered to be a preferred embodiment of the term "comprising of". If hereinafter a group is defined to comprise at least a certain number of embodiments, this is meant to also encompass a group which preferably consists of these embodiments only.

[0026] Furthermore, the terms "(i)", "(ii)", "(iii)" or " (a) ", "(b)", " (c) ", " (d) ", or "first", "second", "third" etc. and the like in the description or in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order.

[0027] It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other sequences than described or illustrated herein. In case the terms relate to steps of a method, procedure or use there is no time or time interval coherence between the steps, i.e. the steps may be carried out simultaneously or there may be time intervals of seconds, minutes, hours, days, weeks etc. between such steps, unless otherwise indicated.

[0028] It is to be understood that this invention is not limited to the particular methodology, protocols etc. described herein as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention that will be limited only by the appended claims.

[0029] The drawings are to be regarded as being schematic representations and elements illustrated in the drawings are not necessarily shown to scale. Rather, the various elements are represented such that their function and general purpose become apparent to a person skilled in the art.

[0030] Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art.

[0031] Independent of the grammatical term usage, individuals with male, female or other gender identities are included within the term.

[0032] As has been set out above, the present invention concerns in one aspect a cartridge for the uptake and analysis of a fluid sample, comprising: (i) an inlet which allows to take up the sample; (ii) a filter zone comprising a filter membrane which allows for separating different components within the sample and for adding reagents to the sample; (iii) a landing zone which allows for delaying the movement of the sample and for blending the sample; (iv) a transport zone which allows for transporting the sample from the landing zone to a reaction chamber, comprising one or more fluidic channels; (v) one or more reaction chambers, which are designed for mixing of reagents with the sample; (vi) one or more reaction chambers, which are designed to capture analytes from the sample and other reactants; (vii) a reaction chamber zone, which is designed for washing, examination and verification of assay reactions; and (viii) one or more stop zones where fluidic flow within the cartridge is stopped.

[0033] The term "cartridge for the uptake and analysis of a fluid sample" as used herein refers to a structure or an instrument, or part of an instrument, or a part of a system, which allows or is suitable for the performance of reactions, in particular molecular reactions involving chemical and / or biological entities and / or particle counting activities and / or the measurement of physical and / or chemical parameters. The cartridge may correspondingly be equipped, for example, with a suitable inlet element, it may comprise one or more surfaces, e.g. reactive surfaces or surfaces with specific functionality, it may comprise, inter alia, and in addition to the zones specifically mentioned herein a washing zone, a mixing zone, a waiting zone, a measurement zone, a waste zone, a reservoir zone, a recollection or a regeneration zone etc. or any sub-portion or combination thereof. The cartridge may further comprise connections between the mentioned elements, e.g. tubes or joints; and / or it may comprise reservoirs and repositories for liquids, fluids, reactants, chemicals, ingredients, samples or any other entity to be used within the cartridge. The zones may further also be equipped with suitable sensor elements allowing the measurement of e.g. light emission, light reflection, light scattering, light interference, temperature changes or temperature gradients.

[0034] The term "inlet which allows to take up the sample" as used herein refers to an opening or tubing structure. The inlet accordingly allows to transport the fluid sample, e.g. in certain embodiments without additional pumping assistance, towards the filter zone (ii) of the cartridge. The inlet may, in certain embodiments, be designed as part of a lid which can be opened and closed and may comprise the inlet in the form of round, rectangular or oval opening. Typically, the inlet is positioned on top or above the filter. In certain embodiments the inlet is designed as three-dimensional structure which is in contact with the filter. It is further envisaged, that the inlet may be in contact with the laminate and / or bottom part of the cartridge, e.g. in order to form a fluidic path preventing the sample to leak away.

[0035] The term "filter zone" as used herein relates to a zone of the cartridge which is designed to separate elements, e.g. cells or particles, from other components of the fluid sample. The filter zone is typically composed of a filter membrane which is located on top of a landing zone.

[0036] The term "filter membrane" as used herein relates to a selective barrier, mainly per-forming the function of a separator, e.g. by allowing for a filtering process. The filter membrane may, accordingly, be designed to allow for partial separation of ingredients of fluid samples such, e.g. blood. Preferably, the filter membrane may be designed to allow for the passage of liquid portions, proteins or subcellular fragments in fluid samples, e.g. of plasma derived from a blood sample. It is further particularly preferred that the filter membrane is incapable of allowing the passage of cellular entities. For example, the filter may impede the passage of blood cells, while allowing the passage of blood plasma.

[0037] In specific embodiments, the filter membrane may comprise one or more chemical or biological reagents. These reagents may be provided in a dried form which are dissolved upon contact with the sample fluid. Thereby the chemical or biological reagent may be contacted with the fluid sample or filtration product, e.g. to modify the pH, stabilize the sample, induce molecular reaction or the like.

[0038] The filter membrane may be composed of any suitable material. Envisaged examples include elastic or foldable polymer materials. Particularly preferred is polyurethane. Also envisaged is the use of micromeshes. In preferred embodiments, these micromeshes may comprise ultrathin fibers, metallic, ceramic or polymeric materials. In certain embodiments, the filter membranes may at least partially be coated with one or more chemical and / or biological agents. The term "coated" as used herein means that all or at least some sectors of the filter membrane material as defined herein is covered by said chemical and / or biological agent(s). The coverage may, in certain embodiments, be present at both sides and / or inside of the filter membrane. In further embodiments, the filter membrane may comprise blocking materials such as bovine serum albumin (BSA), chicken serum albumin (CSA), casein or gelatine. These materials are envisaged as blocking an interaction between specific sample components and the membrane, e.g. a hydrophobic or ionic interaction between sample components such as blood plasma components and polymeric structures of the membrane.

[0039] In preferred embodiments, the filter membrane comprises pores of a diameter of 1.6 µm to 2.8 um. For example, the pores may have a diameter of 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7 or 2.8 um or any value in between the mentioned values. Also envisaged are, in certain specific embodiments, larger or smaller pores. The pores may, in further embodiments, be provided within one membrane with the same diameter, or with two or more different diameters or with multiple pore sizes in one membrane.

[0040] In further embodiments the filter membrane as defined herein covers, for example, at least the area of the body of the structured baseplate as defined herein. In further embodiments, the filter membrane as defined herein partially covers the area of the body of the structured baseplate as defined herein. Accordingly, the filter membrane may preferably contact or essentially contact the landing zone of the cartridge as defined herein.

[0041] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) a landing zone. The term "landing zone" as used herein refers to a structure which allows for a further collection and blending of the sample. The landing zone is a zone which has a filter support area which is located at or comprises sectors of the structured baseplate as defined herein. The filter support area hence corresponds to the space or cavity where the filtered sample components, e.g. blood plasma, first arrives after having passed through the filter being contacted by the elevations of the base plate. The filter support zone thus typically comprises at least the floor or basal part of the structured baseplate and may extend up to the upper section or ceiling, ending at the interface with the filter membrane. The term "filter support" as used herein thus relates to a part of the landing zone which supports the filter membrane. It may comprise, for example, a separate component or may be part of the structured baseplate. It may further also serve to keep distance between the filter membrane and the surface of the baseplate.

[0042] The "structured baseplate" is a component which supports the filter membrane (which is provided on the baseplate) but also serves to keep distance between the filter membrane and the bottom of the baseplate. It accordingly allows for collection of filtered sample fluid. The structured baseplate may have any suitable form and shape. The structure implements the requirements for the provision of a cavity or recess below the filter membrane. In preferred embodiments the baseplate comprises a multitude of elevations. The number of elevations may be adjusted or defined in accordance with the overall design and size of the cartridge, in particular with the overall design and size of the filter zone. For example, the structured baseplate may comprise between 10 and 1500 elevations, e.g. 10, 20, 30, 40, 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 435, 450, 475, 500, 750, 1000, or 1500 or any number in between the mentioned number. Further envisaged are lower or higher numbers. The dimensions of the elevations may be adjusted or defined in accordance with the overall design and size of the cartridge, in particular with the overall design and size of the filter zone. It is preferred that the dimension of the elevations is defined in a manner that the elevations comprise less than 80%, 70%, 60%, 50%, 400, 300, or 200 of the available space of the filter support area, thus allowing for suitable room for mixing and collecting filtered fluid sample components, e.g. blood plasma.

[0043] The elevations may, for example, have any suitable form. Preferably, the elevations are round, oval or rectangular structures. These structures may, for example, have a column-like design. Also envisaged are other design forms which allow for the provision of cavities below the filter membrane. The structured baseplate accordingly conveys the technical effect that filtered fluid, e.g. separated plasma, can mix or be collected prior to the entry into the next zone of the cartridge. In a preferred embodiment the elevation has the form of a pillar structure. Such column-like structures may have any suitable geometric form, e.g. can be round, oval, rectangular, or have complex geometries, e.g. broader foundation or top sections etc.

[0044] According to preferred embodiments the structured baseplate comprises a multitude of contact points with the filter membrane. For example, the structured baseplate may comprise between 10 and 1500 contact points, e.g. 10, 20, 30, 40, 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 435, 450, 475, 500, 750, 1000, or 1500 or any number in between the mentioned number. Such contact points allow for an efficient filtration into the landing zone. Without wishing to be bound by theory, it is assumed that these multiple contact points help to bring fluid from different filter positions to the baseplate. It is further assumed that the space between the filter membrane and the baseplate allows for mixing and flow-through. Thereby a mixing and collection of the filtered fluid sample, e.g. blood plasma, becomes possible.

[0045] The contact points may, for example, be elevations of the baseplate as mentioned herein, which directly interact, i.e. are in direct contact, with the filter membrane.

[0046] In a specific embodiment the structured baseplate comprises a fluidic stop, venting and air system. This system comprises an edge structure which is designed to stop the filtered fluidic sample from flowing to a venting channel. Further, a channel is provided, which allows for air equilibration with the exterior. The channel accordingly connects with the exterior of the cartridge and is designed to allow for air exchange. The term "air" is to be understood in a generic way, thus including any form of gas, e.g. also inert gases or the like.

[0047] In a particularly preferred embodiment the edges of the fluidic stop, venting and air system have a radius of < 0.02 mm. For example, the edges may have a radius of 0.019 mm, 0.018 mm, 0.017 mm, 0.016 mm, 0.015 mm, 0.014 mm, 0.013 mm, 0.012 mm, 0.011 mm, 0.01 mm, 0.009 mm, 0.008 mm, 0.007 mm, 0.006 mm, 0.005 mm, 0.004 mm, 0.003 mm, 0.002 mm or 0.001 mm. Further radius values, e.g. between the mentioned values, are also envisaged.

[0048] In a further embodiment, the structured baseplate comprises a fluidic channel which works as an outlet and connects the baseplate and thus the landing zone to downstream components of the cartridge such as further transport channels, reaction chambers etc. The fluidic channel may, in certain embodiments, have a starting position in the center of the structured baseplate (as, e.g., depicted in Fig. 2, 3 and 4) or, alternatively, at any other position, e.g. at or close to one of the edges of the baseplate. In certain embodiments, the fluidic channel may comprise hydrophilic material as defined herein.

[0049] This landing zone is equipped with components which delay the movement of the filtered fluid sample. In preferred embodiments, the delay of the movement of the filtered fluid sample is achieved by the presence of three-dimensional structures in the landing zone which are coated with hydrophilic material. In certain embodiments, the entire filter support, preferably excluding the edges, may be coated, or a sub-portion thereof, e.g. 900, 80%, 70%, 60%, 50%, 40% or less than 40% or any value in between the mentioned values. In further, specific embodiments the elevations of the structured baseplate may comprise a surface coated with hydrophilic material.

[0050] In a further particularly preferred embodiment, the hydrophilic material is not present at, and does not touch the edges of the filter support. By excluding the edges of the filter support from the presence of hydrophilic material the movement of the filtered fluid sample may be controlled.

[0051] Envisaged examples of hydrophilic material include polysaccharides such as dextran, alginate, agarose, pullulan, or proteins such as albumin, gelatin, collagen, lectin, legumin or vicilline. Also envisaged are polymers such as polyvinyl acetate (PVA), cellulose acetate (CA), or polyacryl nitril (PAN).

[0052] It is particularly preferred that the hydrophilic material is non-biofouling. The term "biofouling" as used herein relates to the spontaneous and unwanted adsorption of biomolecules, cells, or microorganisms on engineered surfaces, being the consequence of the surface properties, such as but not limited to the charge, hydrophobicity, or coating grafting density. Non-biofouling and hydrophilic materials as used in the context of the present invention include, for example, polyethylene glycol (PEG), cyclic peptides or zwitterions such as PTMAO.

[0053] In further preferred embodiments the hydrophilic material, e.g. the non-biofouling material, may also additionally comprise a fluorescence dye. Examples of suitable fluorescence dyes include fluorescein isothiocyanate (FITC), hydroxyfluorescein (FAM), tetrachlorofluorescein (TET), Rhodamine dyes, such as red rhodamine (RBITC), tetramethylrhodamine (TAMRA), rhodamine B (TRITC), Cy series cyanine dyes such as Cy2, Cy3, Cy3B, Cy3.5, Cy5, Cy5.5, Cy7 etc., Alexa series dye such as Alexa Fluor 350, 405, 430, 532, 546, 555, 568, 594, 610, 633, 680, 700, or 750, APDye Fluors or Protein dyes such as phycoerythrin (PE), phycocyanin (PC), or allophycocyanin (APC).

[0054] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) a transport zone. The term "transport zone" as used herein refers to a structure which allows for transporting the filtered fluid sample from the landing zone, where it was delayed and thus blended or mixed, to a further zone, in particular the zone of reaction chambers. This zone is typically composed of capillary channels or tubes which do not require gravity for displacement and allow for a movement of the filtered fluid sample, e.g. blood plasma, without necessity of pumping or assisted movement. The transport zone implements a fluid communication system wherein a filtered fluid sample, or portion thereof or material in the cartridge, can be transported or displaced from landing zone to at least a second zone such as the zone of reaction chambers. It is preferred that the transport zone comprises at least one or more fluidic channels.

[0055] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) one or more reaction chambers, which are designed for mixing of reagents with the sample. The number of mixing reaction chambers may vary depending on the intended use of the cartridge or the assay to be performed. At least one mixing reaction chamber is present. Also envisaged is the presence of two, three, 4, 5, 6 or more mixing reaction chambers. In case the cartridge comprises more than one mixing reaction chamber each of the chambers may comprise different reactants. Also envisaged is a structure, in which the chamber or more than one chamber comprises the same type of reactants. The mixing reaction chambers may be connected directly one to the other, or there may be an individual connection with the transport zone and / or the landing zone as defined herein. The advance of filtered fluid samples into one or more of the mixing reaction chambers may be controlled by the filling state of the chamber. For example, a reaction chamber may not receive further filtered fluid sample if its highest occupancy is reached. The filtered fluid sample may then flow into a second or third etc. mixing chamber.

[0056] The reactants in the mixing reaction chamber may preferably comprise reagents and ingredients to perform a diagnostic analysis such as a buffer, a peptide, a protein, an enzyme, a nucleotide, an antibody, or a dye or any other ingredient necessary for the analysis. The reactants may be provided in a liquid or, preferably, dried or lyophilized form. The analysis may include qualitative and / or quantitative measurements. The analysis is typically based on a biological or biochemical assay format as known to the skilled person. Such assays typically comprise the following general steps: sample processing and manipulation, target specific discrimination or identification, signal or target amplification, signal detection and / or interpretation, and signal enhancement and noise filtering. The assays may include end point assays, e.g. assays in which a single measurement is performed after a fixed incubation period, or kinetic assays, i.e. assays in which measurements are performed multiple times over a fixed time interval. The assays may, for example, include assays in the biological activity of a sample is tested, ligand binding assay, where a ligand binds a receptor, or immunoassays, where the response is an antigen antibody binding type reaction. Also envisaged are assays based on nucleic acids. Further envisaged are enzyme assays, where, for example, enzymes may be tested via their highly repeating activity on a large number of substrates when loss of a substrate or the making of a product may have a measurable attribute like color or absorbance at a particular wavelength or light or electrochemiluminescence or electrical / redox activity. Also contemplated are assays based on radioisotopes, where radiolabeled substrates may be used in radioimmunoassays or equilibrium dialysis assays. Also possible are combinations of any of the above mentioned assays.

[0057] The analysis may, for example, be based on the detection of proteins or protein fragments via binding partners such as antibodies, lectins, peptides or non-immunoglobulin proteins comprising an ankyrin-repeat protein (DARPin), affibody molecule, adnectin, anticalin, affilin, avimer, knottin, fynomer, phylomer, artificial antibody mimic and a kunitz domain peptide. The analysis may, in further embodiments, be based on chemiluminescent signal detection, e.g. based on luminol compounds which can be detected after cupric ions are dissolved from hybrids. Further analysis may include the use of magnetic particles, e.g. comprising one or more of the above-mentioned entities, e.g. antibodies or other interactors, such as magnetic particles. Also envisaged are analysis techniques based on fluorescence in situ hybridization or ELISA techniques. Furthermore, the analysis may be based on microarray structures, which may, for example be provided in one or more specific reaction chambers. Reagents, ingredients, buffers etc. useful for the performance of an assays as mentioned above may, for example, be provided to the device during the manufacturing process, as defined herein below, e.g. by filling in the reagents. Furthermore, in specific embodiments, such reagents, e.g. antibodies, nucleic acids, enzymes, nanoparticles etc. may be immobilized in certain reaction chambers as defined herein. The cartridge may further allow the presence of different types of ingredients, e.g. liquid ingredients, solid ingredients or gaseous ingredients.

[0058] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) one or more reaction chambers, which are designed to capture analytes from the sample and other reactants. The number of capturing reaction chambers may vary depending on the intended use of the cartridge or the assay to be performed. At least one capturing reaction chamber is present. Also envisaged is the presence of two, three, 4, 5, 6 or more capturing reaction chambers. In case the cartridge comprises more than one capturing reaction chamber each of the chambers may comprise different reactants. Also envisaged is a structure, in which the chamber or more than one chamber comprises the same type of reactants. The capturing reaction chambers may be connected directly one the other, or there may be a connection with the transport zone as defined herein. Reactants and structures which allow for the capture of an analyte from the filtered fluid sample may comprise, for example, antibodies, nucleic acids, enzymes, lectins, inorganic catalyst, nanoparticles etc. immobilized at the bottom or walls of the reaction chambers. Also envisaged is the presence of one or more magnetic zones which allow the capture of, e.g., magnetic particles. Such particles may, for example, be bound, selected and / or moved over magnetic zones within the capturing reaction chambers or by the provision of temporary magnetic field. The "other reagents" as used herein may comprise buffers, ingredients etc. which are required for the binding process of the analyte.

[0059] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) a reaction chamber zone, which is designed for washing, examination and verification of assay reactions. This zone may primarily be used to wash and thus removing reactants or other elements which may interfere with measurement process. It preferably further comprises at least one measurement functionality. The term "measurement functionality" as used herein relates to any entity, instrument, unit, compound or composition which allows to determine the status, consistency or quality of a sample; to determine the presence or absence of a specific compound, particle, nucleic acid, protein, structure etc. in a sample; to determine the result of a reaction performed on or with the sample; or to determine any parameter connected to the operation, status, performance, handling etc. of the device itself. Such a measurement functionality may, for example, allow to determine the change of color, or the color homogeneity if a reaction with a dye has been performed. It may, in another example, allow to determine the pH, electric conductivity, turbidity or cloudiness in a solution, or the temperature or any other parameter which is involved in a reaction performed. Thereby the assay results can be obtained and verified, e.g. by additionally performing control or calibration tests.

[0060] In certain embodiments the reaction chamber zone designed for washing, examination and verification may be identical to the zone of one or more reaction chambers, which are designed to capture analytes from the sample. In such embodiments, captured analytes from the sample may be detected in situ, e.g. when bound to the surface of the chamber.

[0061] The cartridge further comprises, as subsequent zone (in the direction of the flow or movement of the filtered fluid sample) one or more stop zones where fluidic flow within the cartridge is stopped. Such stop zones may, for example, be implemented as hydrophobic barriers, which do not allow for the entry or passage of sample fluids.

[0062] In certain embodiments, the cartridge further comprises a container zone which allows for entry and accommodation of filtered fluid sample after having passed the above-mentioned reaction chamber zones. In such a container zone a stop zone as defined above may be present. The container zone may further comprise a backstop which prevents the return of the sample fluid to the other zones.

[0063] In additional, alternative embodiments, the presence of one or more zones as defined above may be optional, e.g. the cartridge may comprise only one or two reaction chambers or reaction chamber zones etc., or one or more other zones are not / only optionally present. The presence of these zones may, in further specific embodiments, be activated, e.g. by the deliberate opening of certain tubes or fluidic channels, e.g. according to need or the detection program to be performed.

[0064] In a further aspect the present invention relates to a system comprising a handheld device capable of operating the cartridge as defined herein and the cartridge as defined herein. The handheld device may, in preferred embodiments, be designed as reader or control unit. It may, accordingly, for example, provide pneumatic and / or electrical connections to the cartridge allowing to induce in the cartridge sample movements, reactions with the sample, measurement of parameters etc. Furthermore, the reader or control functionality of the handheld device may additionally comprise sensors or detecting elements allowing to accumulate measured values in the cartridge. Also, a digital interface, e.g. comprising WiFi and / or Bluetooth and / or WLAN connectivity may be provided within the handheld device, i.e. the system. Obtained measurement results may accordingly be transmitted to remote server structures or cloud servers or be transmitted to further local devices such as desktop computer, tablets or the like.

[0065] In further embodiments, detector elements and assessment units necessary for the detection of the processing state or functioning of the cartridge may be provided by the handheld device. Both elements of the system, i.e. the handheld device and the cartridge, may be connected in a push fit fashion, e.g. as cradle and plug-in module. The handheld device may accordingly be provided with opening or receptacle structures, allowing the connection or introduction of a cartridge. The physical separation of both elements of the system according to the present invention provides the advantage that the same handheld device may be used for multiple analyses, while the cartridge may comprise disposable, non-reusable or non-expensive elements such as chemical reactants or assay components etc. A cartridge according to the present invention is in a preferred embodiment thus envisaged as a single use or disposable product. In a particularly preferred embodiment, the handheld device is an Atellica ®< point-of-care device, e.g. an Atellica ®< VTLi Patient-side Immunoassay Analyzer.

[0066] In a further aspect the present invention relates to a method of producing a cartridge as defined herein comprising the steps: (a) preparing a structured baseplate as defined herein, preferably in a molding process; (b) preparing a filter membrane as defined herein by depositing one or more reagents on the filter membrane; and (c) depositing reagents in zones and chambers as defined herein. In certain embodiments, the method may also comprise the production of other elements of the cartridge as defined herein or shown in Fig. 1, Fig. 2, Fig. 3 or Fig. 4, e.g. a structure comprising an inlet, a filter zone, a landing zone, a transport zone, one or more mixing reaction chambers, one or more capturing reaction chambers, a reaction chamber zone, designed for washing, examination and verification and one or more stop zones.

[0067] The preparation of the structured baseplate or other parts of the cartridge may be performed, for example, according to suitable molding or 3D printing approaches as known to the skilled person. In typical embodiments, the different sections of the cartridge may be produced simultaneously or in a step-by-step manner, e.g. if different materials are used for the production, or if elements of different origin or different consistency are combined. Molding may, for example, be performed as advanced injection molding. Further information would be known the skilled person or can be derived from suitable literature sources such as Czepiel et al., 2023, Materials, 16, 5802.

[0068] A 3D printing procedure is typically performed by specialized 3-D printers which may, in typical embodiments, comprise a controllable printer head and a substrate onto which material can be printed. Examples of suitable 3D printer would be known to the skilled person or can be derived from internet resources such as https: / / www.3dnatives.com or similar sites. The printer head is typically moveable relative to the substrate in a 3 axis motion (i.e. x, y and z axis, where x and y represent the horizontal motion, and z the vertical motion). In typical embodiments, the printer head and / or the substrate are moveable. The term "3D printing" thus relates to the step of delivering material from a printer head to the vicinity of the substrate. In the initial stages of the printing process the material may be delivered onto the substrate surface, thereby forming a layer of material on that surface. Subsequently, the material may be delivered onto a previously deposited layer, thereby forming another layer of material. After the termination of the printing procedure, the printed device can be removed from the substrate. The substrate may, in specific embodiments, be a paper substrate, or form part of the final structure of the device. The printing procedure typically makes use of plastic or metal precursor material to produce the recursive structure elements of the analyzer device of the present invention. Accordingly, the printing procedure may make use, and the printed product accordingly comprise, essentially consists of or consists of thermoplastic, thermosetting plastic, light-activated resin or metal material. In further embodiments, the material of the device, i.e. of the recursive structure elements, may be derived from a precursor material, which is printable from a 3D printer. Suitable precursor material may further be converted into the material of the device, i.e. the recursive structure elements, after printing, e.g. via light or UV-curing. In further embodiments, the recursive structure elements may be partially provided by a 3D printing procedure and partially provided in a different form, e.g. introduced into a 3D printed scaffold after or during the printing process. Examples of suitable thermoplastic materials include, but are not limited to, acrylonitrile butadiene styrene (ABS), polystyrene, polycarbonate, polypropylene, polyvinyl chloride, polyvinyl alcohol (PVA), polyethylene, high-density polyethylene (HDPE), polylactic acid (PLA) and polyamides such as Nylon, cycloolefin copolymer (COC), cellulose acetate, ethylene-vinyl acetate (EVA), fluoroplastics (PTFE, with FEP, PFA, CTFE, ECTFE, ETFE), acrylic / PVC alloys, polyaryletherketone (PAEK), polybutadiene (PBD), or polybutylene (PB) . Thermosetting plastic materials are typically stronger than thermoplastic materials due to the three-dimensional network of bonds and are thus better suited for high-temperature applications. Examples of suitable thermosetting plastic materials include, but are not limited to polyester resins, polyurethans, polyuria / polyurethane hybrids, vulcanized rubber, bakelit, duroplast, urea-formaldehyd, melamine resin, diallyl-phtalate (DPA), epoxy novolac resin, benzoxazines, or polyimides. Examples of suitable light-activated, e.g. UV-curable, resins include, but are not limited to, light-curing acrylate adhesives, light-curing silicones, light-curing cyanoacrylates, light-curing epoxy resins, and light-curing anaerobic adhesives. Examples of suitable metals include, but are not limited to, titanium, aluminium, cobalt, chrome, nickel, gold, platinum and stainless steel. Also envisaged are metal alloys, in particular low-melting point alloys, such as alloys of aluminium, titanium, cobalt-chrome superalloys, nickel superalloys, alloys of steel. Further envisaged are composite materials comprising plastics as defined above with metal additives. Further examples of suitable materials would be known to the skilled person or can be derived from suitable literature sources such as Prabhakar et al., 2020, Materials Today: Proceedings, 45(2).

[0069] In further embodiments, the method of producing a cartridge as defined herein comprises a step of dispensing material on top of a preformed baseplate. Also envisaged is the addition of a further component, e.g. a thin component, between the membrane filter and the baseplate as defined herein.

[0070] In further production steps, a filter membrane as defined herein may be modified by depositing one or more reagents on the filter membrane. Accordingly, chemical or biological reagent may be deposited which are capable of modifying or stabilizing the pH, e.g. buffers, stabilize the sample, e.g. EDTA or induce molecular reactions. These reagents may be provided in a liquid form to the filter membrane and subsequently be dried.

[0071] Similarly, one or more reagents or chemical or biological entities may be deposited in zones and chambers of the cartridge. For example, in the reaction chambers as defined herein reagents or chemical or biological compounds as defined herein may be deposited. For example, said reagents may be provided in a liquid form to the chambers and subsequently be dried.

[0072] In specific embodiments, a 3D production process, which produces the entire cartridge may be suitable paused or interrupted in order to provide the reagents or compounds to the filter or chambers as long as these structures are accessible.

[0073] In a further aspect the present invention relates to a method for uptaking and analyzing a fluid sample in a cartridge as defined herein, comprising at least the steps of: (a) entering a sample to the cartridge; (b) blocking the further penetration at a landing zone; (c) analyzing the sample in a reaction chamber zone; and (d) registering values measured during analysis for subsequent analysis or diagnostic steps. The entering step may, for example, be performed after in a patient or subject the skin, e.g. of a finger, has been perforated to obtain a drop of blood. Alternatively, a fluid sample of a different type, e.g. obtained on a different way, may be entered to the cartridge, e.g. with a pipette or the like. The fluid sample, e.g. blood, may subsequently be delayed in its movement to an analytic site by temporarily blocking its penetration at the landing site. This is achieved by the use of hydrophilic compounds and sharp edges of the filter support as described herein. In a further step the sample is analyzed with respect to the presence and / or concentration of one or more analytes. Such analytes may be small organic or inorganic molecules or proteins / peptides or other compounds. The analysis may be performed as outlined above, e.g. by immunological detection. In a further step the analysis may be finished with the detection of signal corresponding to the presence or absence or concentration of an analyte. For example, the production of light signal, change of pH or the like may be registered as signal. The registered values may subsequently be processed, e.g. in a computer system and be compared with control or reference values to allow for a diagnostic conclusion.

[0074] The term "fluid sample" as used herein relates to any inorganic liquid sample or organic liquid sample. Preferably, the sample is a biological sample. The term "biological sample" as used herein refers to any specimen obtained from a biological organism, preferably a living organism. The term relates also to specimen obtained from non-living, i.e. dead biological organisms, in particular recently deceased organisms. The term "biological organism" includes in general eukaryotic systems and may also comprise sub-portions of eukaryotic systems. In particular, such organisms include higher eukaryotes. In preferred embodiments of the present invention a biological sample may be derived from an animal, preferably from a mammal, e.g. from a cat, a dog, a swine, a horse, a cattle, a sheep, a goat, a rabbit, a rat, a mouse, a monkey. Particularly preferred is a sample obtained from a human being. The sample may be obtained via suitable methods known to the person skilled in the art. The sample used in the context of the present invention should preferably be collected in a clinically acceptable manner, more preferably in a way that nucleic acids (in particular RNA) or proteins are preserved.

[0075] The sample may include body fluids, such as blood, sweat, sputum or saliva, semen and urine. Particularly preferred are body fluid samples such as whole blood, blood plasma, sputum and urine.

[0076] It is particularly preferred that the fluid sample is a blood or blood plasma sample.

[0077] In yet another aspect the present invention relates to the use of the cartridge as defined herein or the system as defined herein for molecular diagnosis, biological sample analysis, preferably blood sample analysis, pharmacology, biological or chemical sample analysis, food analysis, environmental, veterinarian or forensic analysis. The diagnostic or analytical procedures may comprise, in certain embodiments, the provision of the analytical conclusion or results in a corresponding report. Such a report may be provided in any suitable manner or form, e.g. as electronic file, as electronic file distributed or accessible over the internet, e.g. provided in a cloud or deposited on a server, or web-based, e.g. provided on a suitable website. Alternatively, the report may be provided in paper form. The report may be provided and thus drafted in a corresponding form, to a subject or another person associated with the subject, a scientist or group of scientists, a judicial authority, a hospital or clinic, a medical practitioner, or a government office (including information relevant for these entities). The report may accordingly be redacted, modified, extended or adjusted to the above specified recipient.

[0078] The following figures are provided for illustrative purposes. It is thus understood that the figures are not to be construed as limiting. The skilled person in the art will clearly be able to envisage further modifications of the principles laid out herein.

Claims

1. A cartridge for the uptake and analysis of a fluid sample, comprising: (i) an inlet which allows to take up the sample; (ii) a filter zone comprising a filter membrane which allows for separating different components within the sample and for adding reagents to the sample; (iii) a landing zone which allows for delaying the movement of the sample and for blending the sample; (iv) a transport zone which allows for transporting the sample from the landing zone to a reaction chamber, comprising one or more fluidic channels; (v) one or more reaction chambers, which are designed for mixing of reagents with the sample; (vi) one or more reaction chambers, which are designed to capture analytes from the sample and other reactants; (vii) a reaction chamber zone, which is designed for washing, examination and verification of assay reactions; and (viii) one or more stop zones where fluidic flow within the cartridge is stopped.

2. The cartridge of claim 1, wherein the landing zone comprises a structured baseplate with a fluidic channel that is designed to collect and transport filtered sample material, wherein said structured baseplate is arranged below the filter membrane.

3. The cartridge of claim 2, wherein the structured baseplate comprises a multitude of elevations.

4. The cartridge of claim 3, wherein said elevations are formed as round or rectangular structures, preferably as pillar structures.

5. The cartridge of any one of claims 2 to 4, wherein the structured baseplate comprises a multitude of contact points with the filter membrane, allowing for an efficient filtration into the landing zone.

6. The cartridge of any one of claims 2 to 5, wherein the structured baseplate comprises a fluidic stop, venting and air system.

7. The cartridge of claim 6, wherein the fluidic stop, venting and air system comprises edges in the baseplate material, wherein said edges have a radius of < 0.02 mm.

8. The cartridge of any one of claims 1 to 7, wherein the filter membrane comprises pores of a diameter of 1.6 to 2.8 µm.

9. The cartridge of any one of claims 1 to 7, wherein the landing zone comprises a surface coated with a hydrophilic material and / or wherein said elevations of the structured baseplate comprise a surface coated with hydrophilic material.

10. The cartridge of claim 9, wherein the hydrophilic material is non-biofouling.

11. The cartridge of claim 10, wherein the non-biofouling material additionally comprises a fluorescence dye.

12. The cartridge of any one of claims 9 to 11, wherein the hydrophilic material does not touch the edges in the baseplate material.

13. A system comprising a handheld device capable of operating the cartridge of any one of claims 1 to 12 and the cartridge as defined in any one of claims 1 to 12.

14. A method of producing a cartridge as defined in any one of claims 1 to 12, comprising the steps: (a) preparing a structured baseplate as defined in claims 2 to 7, preferably in a molding process; (b) preparing a filter membrane as defined in claim 8 by depositing one or more reagents on the filter membrane; and (c) depositing reagents in zones and chambers as defined in claim 1.

15. A method for uptaking and analyzing a fluid sample in a cartridge as defined in any one of claims 1 to 12, comprising at least the steps of: (a) entering a sample to the cartridge; (b) blocking the further penetration at a landing zone; (c) analyzing the sample in a reaction chamber zone; and (d) registering values measured during analysis for subsequent analysis or diagnostic steps.

16. The cartridge of any one of claims 1 to 12 or the method of claim 15, wherein the fluid sample is a blood or blood plasma sample.

17. Use of the cartridge of any one of claims 1 to 12 or the system of claim 13 for molecular diagnosis, biological sample analysis, preferably blood sample analysis, pharmacology, biological or chemical sample analysis, food analysis, environmental, veterinarian or forensic analysis.

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