Improved analyte detection platform and workflow

Abstract

An analyte detection system or device for high sample throughput, the system or device including a sample holding means for holding a plurality of separate liquid samples for analysis, and an electrochemical aptamer-based sensor for detecting a target analyte in each of the plurality of separate liquid samples for analysis, wherein the separate liquid samples are contacted with the electrochemical aptamer-based sensors for analysis, and a power source and electrical conduits carry a current output from the electrochemical aptamer-based sensors.

Description

[Technical Field]

[0001] The present invention relates generally to the in vitro detection of analytes in clinical samples, including the collection of biological fluids from subjects for testing in a laboratory. More particularly, the present invention relates to improvements in hardware used in in vitro diagnostics and the resulting improvements in workflow. [Background technology]

[0002] Laboratory-based analyte detection methods are undoubtedly an essential part of modern medicine. These methods provide clinicians with important information about their patients to assist in diagnosing new medical conditions, managing existing conditions, assisting in prognosis, or otherwise providing clinically relevant information.

[0003] In many cases, the detection method determines the level of an analyte in a blood sample taken from the patient. Exemplary analytes that can be detected in blood include glucose and hemoglobin A1c (for diabetes), calcium (for kidney disease), troponin and cholesterol / lipids (for heart disease), prostate-specific antigen (for prostate cancer), D-dimer (for coagulation disorders), and C-reactive protein (for infectious and inflammatory diseases).

[0004] Fluids other than blood may be used as samples for analytical methods, including urine, saliva, cell and tissue lysates, bronchoalveolar lavage fluid, cerebrospinal fluid and semen.

[0005] Some analyte detection methods are not performed with a disease state in mind, and include methods for detecting reproductive hormones during pregnancy, antibodies that indicate seroconversion after administration of a vaccine, and dietary factors such as folate and vitamin D.

[0006] Analytes that are exogenous to the patient may also be the subject of detection methods. Common analytes in this regard include viral and bacterial proteins, drugs (prescription and illicit), environmental toxins, etc.

[0007] Typically, the first step is to obtain a clinical sample at the bedside or clinic, where the clinician orders one or more diagnostic methods to be performed on the sample. The sample is given a unique identifier and transported to a competent laboratory for assay of the relevant analytes.

[0008] Today's state-of-the-art laboratories are essentially automated facilities that rely on robotic means to process clinical specimens. Clinical samples typically pass through multiple robotic workstations to perform processes such as sample preparation, reagent addition, reagent removal, and test output acquisition. These processes include numerous liquid transfer steps (often by automated pipetting), incubation steps, washing steps, absorbance or photometric measurement steps, sample movement from one workstation to another, etc.

[0009] Each step typically requires specialized equipment and reagents, which significantly increases the cost of laboratory investigations. Each step also introduces the possibility of error, potentially leading to erroneous information being provided to clinicians. Furthermore, the need for multiple steps results in significant delays in providing results.

[0010] An example of a medium-complexity laboratory-based test is found in the enzyme immunoassay (EIA) for antibodies to hepatitis B surface antigen (anti-HBs) in serum. This EIA is a solid-phase assay that relies on the "sandwich principle" to detect antibodies. A summarized version of the relevant protocol follows:

[0011] Polystyrene beads coated with human hepatitis B surface antigen (HBsAg) are incubated with either a patient specimen or an appropriate control. During the incubation, antibodies, if present, are immunologically bound to the solid-phase antigen. After removal of unbound material and washing of the beads, biotin-labeled human HBsAg (B-HBsAg) and rabbit anti-biotin conjugated to horseradish peroxidase (anti-H-HRPO) are incubated with the antibody-antigen complex on the beads. The biotinylated surface antigen binds to this complex, forming an antigen-antibody-antigen "sandwich." Anti-biotin horseradish peroxidase binds to the biotin component of the "sandwich," forming a solid-phase network. Unbound conjugates are removed, and the beads are washed. Next, a solution of o-phenylenediamine (OPD) containing hydrogen peroxide is added to the beads. After incubation, a yellow color develops in proportion to the amount of anti-HBs bound to the beads. Within limits, the greater the amount of antibody in the sample, the higher the absorbance. The enzymatic reaction is stopped by the addition of acid. The absorbance of the control and sample is determined using a spectrophotometer with the wavelength set at 492 nm.

[0012] As will be readily appreciated, even the above summarized outline demonstrates the need for numerous reagents, multiple items of equipment, many dedicated steps, and extended time to complete. The actual step-by-step protocol for the assay is far more complex and will prove to be even more complicated.

[0013] High-throughput analytical laboratories consist of rooms full of equipment, each with associated purchase / leasing, operating, maintenance, repair, and labor costs. Purchasing and storing reagents, cleaning solutions, and other consumables incurs additional costs. Therefore, such facilities are very expensive to set up and operate, and the associated costs are passed on to relevant government health departments, health insurance companies, or consumers / patients.

[0014] A further problem in analytical laboratories is the lack of flexibility in established workflows. As an example, a subject sample may require analysis for three analytes; in that case, the sample is divided into three aliquots, and each aliquot is batched with other samples for the same test. In some cases, to ensure results are provided in a timely manner, test batches must be run with less than the maximum number of available sample spaces. Running tests on less than the full volume of the sample can waste reagent and equipment resources.

[0015] One aspect of the present invention is to provide an improvement over prior art laboratory-based analyte detection methods and devices. A further aspect of the present invention is to provide a useful alternative to prior art laboratory-based analyte detection hardware and methods.

[0016] The discussion of documents, acts, materials, devices, articles and the like is included in this specification solely for the purpose of providing a context for the present invention. No suggestion or representation is made that any or all of these matters formed part of the prior art or were common general knowledge in the art relevant to the present invention as they existed before the priority date of each claim in this application. Summary of the Invention

[0017] In a first aspect, but not necessarily in its broadest aspect, the present invention provides an analyte detection system or device configured for high sample throughput, the system or device comprising: a sample holding means configured to hold a plurality of separate liquid samples for analysis; one or more electrochemical aptamer-based (EAB) sensors configured to detect a target analyte in each of a plurality of separate liquid samples for analysis; contacting means configured to contact each of a plurality of separate liquid samples for analysis with one or more EAB sensors; a power source configured to provide power to the one or more EAB sensors; one or more electrical conduits configured to carry a current output from the one or more EAB sensors; Equipped with.

[0018] In one embodiment of the first aspect, each of the multiple separate liquid samples for analysis is contacted with a dedicated EAB.

[0019] In one embodiment of the first aspect, the holding means is a plurality of collocated vessels.

[0020] In one embodiment of the first aspect, the holding means is a multi-well plate or a rack of vessels.

[0021] In one embodiment of the first aspect, the multiwell plate comprises at least 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48 or 96 wells.

[0022] In one embodiment of the first aspect, the rack of vessels comprises at least 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48 or 96 vessels.

[0023] In one embodiment of the first aspect, the contacting means is a pipette having a lumen, the system or device is configured to aspirate a liquid sample for analysis into the lumen, and the one or more EAB sensors are disposed within the lumen.

[0024] In one embodiment of the first aspect, each of the plurality of separate liquid samples for analysis has a dedicated pipette.

[0025] In one embodiment of the first aspect, the pipette includes (i) one or more electrical conduits for connecting a power source to the one or more EAB sensors, and (ii) one or more electrical conduits configured to carry an output current from the one or more EAB sensors.

[0026] In one embodiment of the first aspect, the system or device comprises a pipette mount, the pipette being attachable to and detachable from the pipette mount.

[0027] In one embodiment of the first aspect, the pipette mount includes an electrical connector configured to connect to (i) one or more electrical conduits of the pipette for connecting a power source to the one or more EAB sensors, and (ii) one or more electrical conduits configured to carry output current from the one or more EAB sensors.

[0028] In one embodiment of the first aspect, the system or device comprises a liquid aspirating means configured to aspirate a liquid sample from outside the pipette into the lumen of the pipette.

[0029] In one embodiment of the first aspect, the contacting means is a sample retaining means and the one or more EAB sensors are positioned within or around the sample retaining means so as to contact the liquid sample retained therein.

[0030] In one embodiment of the first aspect, the sample holding means includes (i) one or more electrical conduits for connecting a power source to the one or more EAB sensors, and (ii) one or more electrical conduits configured to carry output current from the one or more EAB sensors.

[0031] In one embodiment of the first aspect, the system or apparatus comprises a sample holder mount, the sample holder being attachable to and detachable from the sample holder mount.

[0032] In one embodiment of the first aspect, the sample holder mount includes an electrical connector configured to connect to (i) one or more electrical conduits of the sample holder for connecting a power source to the one or more EAB sensors, and (ii) one or more electrical conduits configured to carry output current from the one or more EAB sensors.

[0033] In one embodiment of the first aspect, each of the multiple separate liquid samples for analysis is contacted with a dedicated EAB.

[0034] In one embodiment of the first aspect, the one or more EAB sensors are in the form of one or more biosensor probes that are contactable with each of a plurality of separate liquid samples for analysis.

[0035] In one embodiment of the first aspect, the contacting means is a transferring means configured to bring together one or more biosensor probes and a plurality of separate liquid samples for analysis.

[0036] In one embodiment of the first aspect, the moving means comprises a motorized mechanism configured to lower the one or more biosensor probes towards the holding means so as to contact the plurality of separate liquid samples for analysis, and / or to raise the holding means towards the one or more biosensor probes so as to contact the plurality of separate liquid samples for analysis.

[0037] In one embodiment of the first aspect, each of the multiple separate liquid samples for analysis is contacted with a dedicated EAB.

[0038] In one embodiment of the first aspect, the system or device includes a processor configured to receive current values ​​or derivative values ​​thereof output by one or more EAB sensors and convert the current values ​​or derivative values ​​thereof into clinically relevant values.

[0039] In one embodiment of the first aspect, the clinically relevant value is the amount or concentration of the target analyte.

[0040] In one embodiment of the first aspect, the system includes program instructions stored in an electronic memory, the program instructions configured to convert the current value or a derivative thereof into a clinically relevant value.

[0041] In one embodiment of the first aspect, the system comprises a networking module in operative communication with the processor, the networking module in network communication with a server at an analytical laboratory, and the networking module transmits the current value or its derivative value, or a clinically relevant value, to the server in association with the sample identifier.

[0042] In one embodiment of the first aspect, the system or device is configured as a sample processing and analysis robot.

[0043] In one embodiment of the first aspect, the system or device is a substantially self-contained device.

[0044] In one embodiment of the first aspect, the substantially self-contained device is dimensioned for placement on a laboratory bench.

[0045] In a second aspect, the invention includes a collocation of a pipette, a sample vessel, or a probe, each of which includes one or more electrochemical aptamer-based (EAB) sensors configured to detect a target analyte.

[0046] In one embodiment of the second aspect, the pipettes, sample vessels or probes are spatially arranged in an orderly manner so as to be usable by a robotic sample processing and analysis system or device.

[0047] In one embodiment of the second aspect, the regular arrangement is a regular spacing between pipettes, sample vessels or probes.

[0048] In one embodiment of the second aspect, the regular arrangement is an array arrangement or a grid arrangement.

[0049] In one embodiment of the second aspect, the co-location is configured to be operable with the system or apparatus of any embodiment of the first aspect.

[0050] In a third aspect, the present invention provides a method for simultaneously analyzing multiple samples for one or more target analytes, the method comprising: placing each of a plurality of separate liquid samples for analysis in a separate container; contacting each of the separate liquid samples for analysis with one or more electrochemical aptamer-based (EAB) sensors; receiving a current value output from each of the one or more EAB sensors; converting the current value or a derivative thereof to a clinically relevant value; Includes.

[0051] In one embodiment of the third aspect, at least about 2, 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48 or 96 samples are analyzed simultaneously.

[0052] In one embodiment of the third aspect, one or more EAB sensors are disposed in or around a pipette tip configured to aspirate a liquid sample, or in or around a probe configured to contact a liquid sample, or in or around a container configured to hold a liquid sample.

[0053] In one embodiment of the third aspect, the method is at least partly performed by a system or apparatus of any embodiment of the first aspect.

[0054] In one embodiment of the third aspect, the method employs the collocation of any of the embodiments of the second aspect.

[0055] In one embodiment of the third aspect, the method lacks or does not require any one or more of the following steps: sample preparation, dilution, centrifugation, purification, reagent addition, washing, mixing, incubation, heating, cooling, light detection, light spectroscopy, light scattering analysis, light absorption analysis, visual inspection, elution, radiation detection, inter-container sample transfer, and inter-device sample transfer.

[0056] In a fourth aspect, the present invention provides a method of assembling a customized set of pipettes, sample containers or probes capable of detecting one or more analytes in a sample, the method comprising: providing a library comprising a plurality of pipettes, sample containers or probes, each pipette, sample container or probe having an associated EAB sensor capable of detecting a particular analyte; selecting two or more pipettes, sample vessels or probes from an associated library; juxtaposing two or more selected pipettes, sample containers or probes; Includes.

[0057] In a fifth aspect, the present invention provides a method of manufacturing a customized plate comprising a plurality of wells, each well capable of detecting one or more analytes in a sample, the method comprising: providing a library comprising a plurality of electrochemical aptamer-based (EAB) sensors capable of detecting a particular analyte; selecting two or more of the EAB sensors; associating each of the wells with one of the selected EAB sensors; Includes. [Brief explanation of the drawings]

[0058] [Figure 1]1 illustrates a method and apparatus of the present invention for analyzing multiple blood samples from a group of patients, each patient requiring an assay for a different analyte.

[0059] [Figure 2] FIG. 1 illustrates the use of a pipette incorporating EAB sensors to aspirate samples from a microtiter plate. Upon contact with the sample, each EAB sensor is interrogated to determine the analyte concentration.

[0060] [Figure 3] Figure 1 illustrates the use of microtiter plate wells incorporating EAB sensors to receive samples dispensed by a conventional pipette. Upon contact with the sample, each EAB sensor is interrogated to determine the analyte concentration.

[0061] [Figure 4] 1 illustrates the use of tubes installed in a rack, each with an integrated EAB sensor. Sample is dispensed into the tubes with a regular pipette, and upon contact with the sample, each EAB sensor is interrogated to determine the analyte concentration.

[0062] [Figure 5] FIG. 1 illustrates the use of EAB sensor-coated probes to contact samples held in the wells of a microtiter plate. Upon contact with the sample, each EAB sensor is interrogated to determine the concentration of an analyte.

[0063] [Figure 6] 1 illustrates the generation of a customized pipette set from a pipette library, where the customized set includes a combination of pipette types, each capable of detecting a specific analyte with an associated EAB sensor.

[0064] [Figure 7]1 shows the generation of customized probe sets from a probe library. A customized set includes a combination of probe types, each capable of detecting a specific analyte with an associated EAB sensor.

[0065] [Figure 8] 1 illustrates the generation of customized tube sets from a tube library. The customized sets contain a combination of probe types, each capable of detecting a specific analyte with an associated EAB sensor.

[0066] [Figure 9] 1 shows the generation of customized microtiter plates from aptamer libraries. The customized plates contain wells, each capable of detecting a specific analyte by an associated EAB sensor.

[0067] Unless otherwise indicated herein, drawing features labeled with the same numbers when used across different drawings are considered to be the same features or at least functionally similar features.

[0068] The drawings are not made to any particular scale or dimension, and are not intended to be entirely accurate representations of the various embodiments. DETAILED DESCRIPTION OF THE INVENTION

[0069] After considering this description, it will be apparent to one skilled in the art how the present invention may be implemented in various alternative embodiments and applications. However, while various embodiments of the present invention are described herein, it is understood that these embodiments are presented by way of example only, and not limitation. Thus, this description of various alternative embodiments should not be construed as limiting the scope or breadth of the present invention. Furthermore, statements of advantages or other aspects apply to particular example embodiments and not necessarily to all embodiments, or indeed to any embodiment covered by the claims.

[0070] Throughout the description and claims of this specification, the word "comprise" and variations of this word, such as "comprising" and "comprises," are not intended to exclude other additional elements, components, integers or steps.

[0071] References throughout this specification to "one embodiment" or "an embodiment" mean that a particular feature, structure, or characteristic described in connection with that embodiment is included in at least one embodiment of the invention. Thus, the appearances of the phrase "in one embodiment" or "in an embodiment" in various places throughout this specification are not necessarily all referring to the same embodiment, although they may.

[0072] As used herein, terms describing positions such as "lateral," "across," "above," "below," "higher," "lower," "upward," "downward," "plan view," etc., are intended to be considered with respect to the analyzer being used in its normal upright position so that the liquid sample can flow downward under the force of gravity.

[0073] The present invention is based, at least in part, on the inventors' discovery that the use of electrochemical aptamer-based (EAB) sensors in the context of a robotic, high-throughput sample analysis laboratory offers significant advantages over the same operations. For example, an EAB sensor can be associated with a pipette, probe, or sample container, and analyte concentration can be determined in a single contact step with the sample. This contrasts with the lengthy, multi-step testing methods used by prior art high-throughput devices and systems, making the present invention highly suitable for application in analytical laboratories that process very large numbers of samples per day. Furthermore, some versions of the present invention significantly reduce equipment requirements by enabling high-throughput processing using only a single item of equipment. Another potential advantage is that laboratory workflows can be modified to more efficiently process the varying numbers of different tests offered.

[0074] Potentially useful EAB sensors in the context of the present invention can be potentiometric, amperometric, or conductivity types. In potentiometric sensors, local equilibrium is established at the sensor interface, where either the electrode or membrane potential is measured, and information about the sample is derived from the potential difference between the two electrodes. Amperometric sensors rely on a potential applied between a reference electrode and a working electrode to cause the oxidation or reduction of a redox-active species, and the resulting current is measured. Conductivity sensors rely on measuring conductivity over a range of frequencies.

[0075] EAB sensors have been shown to reliably and specifically detect drugs in patient fluids. These types of sensors are typically amperometric, with an aptamer (such as DNA, RNA, or XNA) attached to a working electrode. Gold is often used as the probe surface of the working electrode. The aptamer has an associated redox-active species that acts as a reporter. The redox reporter is often methylene blue. Upon target (drug) binding, the aptamer undergoes a conformational change, bringing the redox reporter closer to the working electrode surface. This increased proximity increases electron transfer from the redox reporter to the electrode. The increased rate of electron transfer contributes to a change in the faradaic current detected by a potentiostat.

[0076] Aptamers are small (usually 20-60 nucleotides) single-stranded RNA, DNA, or XNA oligonucleotides that can bind to target drugs with high affinity and specificity. Aptamers can be thought of as the nucleotide analogue of antibodies, but their production is an in vitro cell-free process that is significantly easier and cheaper than producing antibodies by cell culture or in vivo methods.

[0077] Aptamers are typically available in large numbers (up to 10 18 ) are selected from combinatorial libraries with different oligonucleotides. Although RNA aptamers offer significantly greater structural diversity compared to DNA aptamers, their application is complicated by stability issues in the presence of RNases, high temperatures, and unfavorable pH.

[0078] The selection of aptamers selective for a given drug can be facilitated by a process known as SELEX (systematic evolution of ligands by exponential enrichment). This process can be thought of as two alternating steps. In the first step, library oligonucleotides are amplified to the desired concentration by polymerase chain reaction (PCR). For RNA aptamer selection, single-stranded oligoribonucleotides are generated by in vitro transcription of double-stranded DNA using T7 RNA polymerase. For DNA aptamers, a pool of single-stranded oligodeoxyribonucleotides is generated by strand separation of double-stranded PCR products. In the second step, the products of the amplification are incubated with the target drug, and the oligonucleotides that bind to the drug are used in the next round of SELEX.

[0079] Separation of oligonucleotides with higher affinity for the target drug and removal of unbound oligonucleotides is achieved by intense competition for the binding site. Selection pressure increases with each SELEX round. Maximum enrichment of the oligonucleotide pool with aptamers with the strongest affinity for the target molecule is typically achieved after 5-15 rounds.

[0080] EAB sensors are typically incorporated into a circuit with a reference electrode. The reference electrode is the site of a known chemical reaction with a known redox potential. For example, a reference electrode based on the silver-silver chloride (Ag|AgCl) redox couple has a fixed, known potential that forms the point from which the redox potential of the working electrode is measured. Also typically included in the circuit is a counter electrode, which functions as the cathode or anode relative to the working electrode. Because the applied voltage bias does not pass through the reference electrode (due to the impedance of the potentiostat), any potential developed is attributed to the working electrode. A current is measured as the potential of the interrogation electrode versus the stable potential of the reference electrode. The potential difference generates a current in the circuit, thereby generating an output signal. The signal quantifies target binding by relying on electron transfer, ideally stoichiometrically proportional to target binding.

[0081] The present invention will now be described in more detail with reference to the following non-limiting embodiments.

[0082] Refer to Figure 1, which illustrates an exemplary workflow for using the high-throughput analyte detection device 10. The device 10 is self-contained and can perform a wide range of analyte detection methods without the need for additional equipment. Furthermore, the device 10 detects target analytes through a single step of contacting the sample with the EAB sensor.

[0083] In this exemplary workflow, multiple subjects (15, 20, 25, ... n) each provide a blood sample into a collection tube (30, 35, 40) for analysis of an analyte, such as a drug, metabolite, or antibody of particular specificity. Aliquots of blood from each collection tube (30, 35, 40) are placed into wells of a 96-well microtiter plate (45), one aliquot per well.

[0084] The microtiter plate (45) is mounted in a tray (50) of a high-throughput analyte detection device (10). The device (10) includes an array of pipettes (not visible in FIG. 1 ) arranged in a 12×8 grid and aligned with the wells of the 96-well microtiter plate (45). As explained more fully below, the array of pipettes is initially positioned above the microtiter plate (45) and lowered into the wells of the microtiter plate (45) for analysis, at which point sample from each well is aspirated into the respective pipette lumen where it comes into contact with the EAB sensor.

[0085] After interrogating the EAB sensors, the analyte concentration for each subject (15, 20, 25) is displayed on the screen (55) for review by the operator, if desired.

[0086] The analyte concentrations for each subject (15, 20, 25) are transmitted as an electronic file, data packet, or otherwise to a laboratory server (100) and stored in the associated analytical sample laboratory relational database (105). The analyte concentrations are stored in association with an identifier, such as the associated subject's name (optionally with date of birth) or other unique identifier, such as a health insurance number, social security number, patient number, or the like.

[0087] At the end of the analysis, the microtiter plate (45) is removed from the tray (50) and discarded.

[0088] The pipette tips are removable from the high-throughput analyte detection device (10) and are also discarded.

[0089] In preparation for the next analysis, a new set of pipette tips is placed into the high-throughput analyte detection device (10). The particular set of tips selected depends on the analyte to be detected in the next analysis. For example, if the target analyte is troponin, each pipette in the set will have an aptamer specific for troponin.

[0090] Analytical laboratories may have a "library" of pipettes from which an operator (or even a machine) may select according to the desired analyte. In some situations, pipettes specific to different analytes may be mixed to form a set of pipettes. For example, each sample may be tested for troponin and creatinine phosphokinase, in which case each subject's blood is placed in two wells, with the contents of the first well aspirated into a troponin-sensing pipette and the contents of the second well aspirated into a creatinine phosphokinase-sensing pipette. Alternatively, a single pipette may contain aptamers capable of sensing more than one analyte.

[0091] A mixed set of pipettes may be used when each subject requires analysis for a single target analyte, but two different analyte runs are performed on a single microtiter plate. For example, a first group of subjects may require analysis for glucose, and a second group of subjects may require analysis for testosterone. In this case, the microtiter plate may be divided into half for glucose and the other half for testosterone.

[0092] As will be appreciated from the above, the use of aptamer-loaded pipettes offers significant flexibility in analytical laboratory operations: highly customized pipette sets can be created to allow multiple selected analytes to be assayed on a single microtiter plate.

[0093] As an alternative to plate-based assays, the present invention may be implemented using a rack of tubes. In this situation, the pipette tip is lowered into the tubes rather than into the wells of a plate. When tubes are used, the EAB sensor may be located within the pipette or within the tube itself. A set of tubes may be provided, with each tube selected according to the analyte sensing specificity required for the associated sample. In that sense, tube selection and mixing may be considered similar to that described above for pipettes.

[0094] The use of EAB sensors in laboratory-based diagnostics enables the customization described above. Many prior art assays involve very different analytical methods, which hinders the ability to run different assays on the same plate or rack of tubes. For example, one method may be an enzyme-linked immunosorbent assay involving multiple steps, while another method may rely on magnetic bead technology. The very different process steps prevent prior art assays from being run together. In contrast, aptamer-based detection methods may rely on only a single contacting step, regardless of the target analyte. For example, the contacting step in the exemplary workflow of FIG. 1 is performed by simply aspirating the sample into a pipette so that it contacts the aptamer in the pipette lumen. At that point, the EAB sensor is interrogated by applying a potential (such as by square-wave voltammetry), providing a reliable current output proportional to the amount of analyte present within seconds.

[0095] The use of a single contact step allows for significant time savings that dramatically improve analytical laboratory throughput. Eliminating the need for a wash step allows for the use of multiple reagents at different points in the detection method, and the transfer of samples between items or instruments, allowing for the processing of thousands. In this exemplary workflow, a 96-well microtiter plate can be used to analyze approximately 60, 70, or 80 samples (with multiple wells typically reserved for use as controls, generating standard curves, etc.).

[0096] Referring to Figure 2, Figure 2 shows a portion of the interior of the high-throughput analyte detection device 10 of Figure 1. In particular, pipettes are shown installed in the device 10, three of which are marked (200, 205, 210). In fact, there are second, third, fourth, fifth, sixth, seventh, and eighth rows of pipettes to provide an array of 96 pipettes.

[0097] Each of the pipettes (200, 205, 210) has a different aptamer and therefore each senses a different analyte, so a single microtiter plate passed through the device (10) only once provides results for three different analytes simultaneously.

[0098] As shown in Figure 2, each pipette is mounted on a hollow mount (marked 215). The hollow is in the form of a channel in gas communication with the pipette lumen. The mount (215) is sized to allow the pipette to form a pressure fit with it when pressed in. This fit is tight enough to allow for some air tightness, but not overly tight so as to make the pipette difficult to remove.

[0099] Each of the mounts (215) forms part of the head (220). A hollow channel in each mount (215) is pneumatically connected to a common cavity in the body portion of the head (220), which is in turn pneumatically connected to a pneumatic system (225).

[0100] The embodiment of Figure 2 operates by aspirating sample from each well of the microplate upward and into each pipette. Aspiration is achieved by a pneumatic system (225) establishing a weak to moderate vacuum within the head (220), which, via the mount (215), establishes a vacuum in the pipette lumen to draw the sample into it.

[0101] The embodiment of Figure 3 is an alternative configuration in which each well of a microtiter plate (45) has an EAB sensor therein. A pipette dispenses sample into the well by generating positive pressure within the pipette lumen using a pneumatic system (225), and an interrogation current is applied to provide an output, similar to the embodiment of Figure 2. This embodiment offers the advantage of requiring a single contact step to obtain a result, but is less flexible than the embodiment of Figure 2. In particular, note that because each well is physically coupled together in the form of a plate, it is not possible to freely mix different EAB sensors to form a customized set. Thus, while a plate can contain wells each having a different EAB sensor disposed therein, the type of EAB sensor must be determined at the time the plate is manufactured and cannot be changed.

[0102] This drawback of the embodiment of Figure 3 is overcome by the embodiment of Figure 4, which has multiple tubes (three marked 230, 235, 240) held in a grid arrangement by a rack (245). Each of the tubes (230, 235, 240) may contain a different EAB sensor. As with the pipette of Figure 2, the tubes (230, 235, 240) may be freely selected and mixed to provide a rack with a customized set of tubes.

[0103] Referring now to the embodiment of FIG. 5, the embodiment of FIG. 5 is a variation of the embodiment of FIG. 2, whereby the pipette is replaced with probe members (three marked 260, 265, and 270), each coated at its end with a different type of EAB sensor. The probes (260, 265, and 270) are interchangeable and pressure-fit into a mount (marked 275). The probes (260, 265, and 270) are lowered into the sample by a motorized transport arrangement, and a test potential is applied to each of the probes (260, 265, and 270). This embodiment shares the advantage of the embodiment of FIG. 2 in that customized probe sets can be provided by selecting and mixing probes as needed.

[0104] The present invention is suitable for computer implementation, given that the output of the EAB sensor is an electrical signal that can be stored electronically as a numerical value (e.g., a current value) in volatile memory and manipulated and analyzed by an associated processor under the direction of software.

[0105] As will be appreciated by those skilled in the art, the present invention may be partially or wholly deployed via one or more processors that execute computer software, program code, and / or instructions on the processors. The processor may be part of a server, client, network infrastructure, mobile computing platform, fixed computing platform, or other computing platform. The processor may be any type of computational or processing device capable of executing program instructions, code, binary instructions, etc. The processor may be or include a signal processor, digital processor, embedded processor, microprocessor, or any variant such as a coprocessor (mathematical coprocessor, graphic coprocessor, communication coprocessor, etc.) and the like that may directly or indirectly facilitate the execution of program code or program instructions stored thereon.

[0106] Additionally, the processor may allow for the execution of multiple programs, threads and codes.

[0107] Threads may be executed simultaneously to improve processor performance and facilitate simultaneous operation of applications. In implementations, the methods, program code, program instructions, etc. described herein may be implemented in one or more threads. Threads may spawn other threads with associated priorities, and the processor may execute these threads based on priority or any other order based on instructions provided in the program code. The processor may include memory for storing the methods, code, instructions, and programs as described herein and elsewhere.

[0108] Any processor or mobile device or server may access, via an interface, a storage medium that may store methods, codes, and instructions as described herein or elsewhere. Storage media associated with a processor for storing methods, programs, codes, program instructions, or other types of instructions executable by a computing or processing device may include solid-state memory and hard disk memory.

[0109] A processor may include one or more cores, which may increase the speed and performance of a multiprocessor. In some embodiments, a processor may be a dual-core processor, a quad-core processor, or other chip-level multiprocessor that combines two or more independent cores (called a die).

[0110] The methods and systems described herein may be deployed, in part or in whole, via one or more hardware components executing software on a server, client, firewall, gateway, hub, router, or other such computer and / or networking hardware. Software programs may be associated with a server, which may include a file server, print server, domain server, Internet server, intranet server, and other variations such as secondary servers, host servers, distributed servers, etc. A server may include one or more of memory, a processor, computer-readable media, storage media, ports (physical and virtual), communication devices, and interfaces capable of accessing other servers, clients, computers, and devices via wired or wireless media, etc. The methods, programs, or codes described herein and elsewhere may be executed by a server. Additionally, other devices required for the execution of methods as described in this application may be considered part of the infrastructure associated with the server.

[0111] A server may provide an interface to other devices, including, but not limited to, clients, other servers, printers, database servers, print servers, file servers, communication servers, distributed servers, etc. Furthermore, this coupling and / or connection may facilitate remote execution of programs across a network. Networking some or all of these devices may facilitate parallel processing of a program or method at one or more locations without departing from the scope of the present invention. Additionally, any of the devices connected to a server via an interface may include at least one storage medium capable of storing methods, programs, code, and / or instructions. A central repository may provide program instructions to be executed on different devices. In this implementation, the remote repositories may serve as storage media for program code, instructions, and programs.

[0112] A software program may be associated with a client, which may include a file client, a print client, a domain client, an Internet client, an intranet client, and other variations such as a secondary client, a host client, a distributed client, etc. A client may include one or more of: one or more memories, processors, computer-readable media, storage media, ports (physical and virtual), communication devices, and interfaces that can access other clients, servers, computers, and devices via wired or wireless media, etc. The methods, programs, or code described herein and elsewhere may be executed by a client. Additionally, other devices required for the execution of methods as described in this application may be considered part of the infrastructure associated with the client.

[0113] A client may provide an interface to other devices, including, but not limited to, servers, other clients, printers, database servers, print servers, file servers, communication servers, distributed servers, etc. Furthermore, this coupling and / or connection may facilitate remote execution of programs across a network. Networking some or all of these devices may facilitate parallel processing of a program or method at one or more locations without departing from the scope of the present invention. Additionally, any of the devices connected to a client via an interface may include at least one storage medium capable of storing methods, programs, code, and / or instructions. A central repository may provide program instructions to be executed on different devices. In this implementation, remote repositories may serve as storage media for program code, instructions, and programs.

[0114] The methods and systems described herein may be deployed, in part or in whole, over a network infrastructure. The network infrastructure may include elements such as computing devices, servers, routers, hubs, firewalls, clients, personal computers, communication devices, routing devices, and other active and passive devices, modules, and / or components known in the art. Computing and / or non-computing devices associated with the network infrastructure may include storage media, apart from other components. The processes, methods, program codes, and instructions described herein and elsewhere may be executed by one or more of the network infrastructure elements.

[0115] The methods, program codes, calculations, algorithms, and instructions described herein may be implemented on a cellular network having multiple cells. The cellular network may include mobile devices, cell sites, base stations, repeaters, antennas, towers, etc. The cell network may be a GSM, GPRS, 3G, 4G, 5G, EVDO, mesh, or other network type.

[0116] The methods, program codes, calculations, algorithms, and instructions described herein may be implemented on or via a mobile device. Mobile devices may include mobile phones, mobile personal digital assistants, laptops, palmtops, netbooks, pagers, e-readers, etc. These devices may include, among other components, storage media such as flash memory, buffers, RAM, ROM, and one or more computing devices. The computing devices associated with the mobile devices may be enabled to execute program codes, methods, and instructions stored thereon.

[0117] Alternatively, the mobile device may be configured to execute instructions in cooperation with other devices. The mobile device may communicate with a base station that interfaces with a server and is configured to execute program code. The mobile device may communicate over a peer-to-peer network, a mesh network, or other communication network. The program code may be stored in a storage medium associated with the server and executed by a computing device embedded within the server. The base station may include a computing device and a storage medium. The storage device may store program code and instructions executed by a computing device associated with the base station.

[0118] Computer software, program code and / or instructions may be stored on and / or accessed from computer-readable media, which may include computer components, devices, and recording media that hold digital data used in computations for a period of time, storage known as random access memory (RAM), and mass storage devices, typically for more permanent storage, such as forms of magnetic storage like optical disks and hard disks.

[0119] The methods and systems described herein may transform physical and / or intangible items from one state to another. The methods and systems described herein may also transform data representing physical and / or intangible items from one state to another.

[0120] The elements described and illustrated herein may imply logical boundaries between the elements. However, according to software or hardware engineering practices, the illustrated elements and their functionality may be implemented on a computer via a computer-executable medium having a processor capable of executing program instructions stored thereon, as a monolithic software structure, as a stand-alone software module, or as a module using external routines, code, services, etc., or any combination thereof; all such implementations may be within the scope of the present disclosure.

[0121] Additionally, the illustrated elements may be implemented on machines capable of executing program instructions. Thus, while the present description describes functional aspects of the disclosed systems, the specific configuration of software for implementing those functional aspects should not be inferred from these descriptions unless explicitly stated or apparent from the context. Similarly, it will be understood that the various steps identified and described above may be varied, and the order of steps may be adapted to particular applications of the techniques described herein. All such variations and modifications are intended to fall within the scope of the present disclosure. Thus, the illustration and / or description of the order of various steps should not be understood as requiring a particular order of performance of those steps unless required by a particular application or unless explicitly stated or apparent from the context.

[0122] The methods and / or processes described above, and steps thereof, may be implemented in hardware, software, or any combination of hardware and software suitable for a particular application. Hardware may include general-purpose computers and / or special-purpose computing devices or specific computing devices or specific aspects or components of specific computing devices. The processes may also be implemented in one or more microprocessors, microcontrollers, embedded microcontrollers, programmable digital signal processors, or other programmable devices, along with internal and / or external memory. The processes may also, or instead, be embodied in application-specific integrated circuits, programmable gate arrays, programmable array logic, or any other device or combination of devices that can be configured to process electronic signals. It will further be understood that one or more of the processes may be implemented as computer-executable code executable on a computer-readable medium.

[0123] Application software may be written using a structured programming language such as C, an object-oriented programming language such as C++, or any other high-level or low-level programming language (including assembly language, hardware description languages, and database programming languages ​​and techniques) that can be stored, compiled, or interpreted for execution on one of the above devices, as well as heterogeneous combinations of processors, processor architectures, or combinations of different hardware and software, or any other machine capable of executing program instructions.

[0124] Thus, in one aspect, each of the methods and combinations thereof described above may be embodied in computer-executable code that performs the steps when executed on one or more computing devices. In another aspect, the methods may be embodied in a system that performs the steps, may be distributed in some manner across multiple devices, or all of the functionality may be integrated into a dedicated stand-alone device or other hardware. In another aspect, the means for performing the steps associated with the processes described above may include any of the hardware and / or software described above. All such permutations and combinations are intended to fall within the scope of the present disclosure.

[0125] Any of the methods disclosed herein may be performed by application software executable on any past, present, or future operating system of a processor-enabled device, such as Windows®, Linux®, Android®, iOS®, etc. It will be understood that any software may be distributed across several devices or in a "software-as-a-service" or "platform-as-a-service" format, whereby participants need only some computer-based means of engaging with the software.

[0126] The present invention has been described primarily with reference to the analysis of clinical samples taken from subjects, particularly human subjects. It will be understood that the present invention is applicable to other applications requiring high-throughput testing of samples for one or more analytes, including, but not limited to, veterinary medicine, agriculture, scientific research, quality control and quality assurance in manufacturing environments, food safety, and the analysis of environmental samples such as water and soil. Those skilled in the art will appreciate that the invention described herein is susceptible to further variations and modifications other than those specifically described. It will be understood that the present invention includes all such variations and modifications that are within the spirit and scope of the present invention.

[0127] Accordingly, the spirit and scope of the present invention should not be limited by the foregoing examples, but should be understood in the broadest sense allowed by law.

Claims

1. 1. An analyte detection system or device configured for high sample throughput, the system or device comprising: a sample holding means configured to hold a plurality of separate liquid samples for analysis; one or more electrochemical aptamer-based sensors configured to detect a target analyte in each of the plurality of separate liquid samples for analysis; contacting means configured to contact each of the plurality of distinct liquid samples for analysis with the one or more electrochemical aptamer-based sensors; a power source configured to provide power to the one or more electrochemical aptamer-based sensors; one or more electrical conduits configured to carry a current output from the one or more electrochemical aptamer-based sensors; 1. An analyte detection system or device comprising:

2. each of said plurality of distinct liquid samples for analysis is contacted with a dedicated electrochemical aptamer-based sensor; 10. The system or device of claim 1.

3. the holding means being a plurality of juxtaposed containers; 3. A system or apparatus according to claim 1 or 2.

4. The holding means is a multi-well plate or a container rack; A system or device according to any one of claims 1 to 3.

5. The multiwell plate comprises at least 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48, or 96 wells; A system or device according to any one of claims 1 to 4.

6. The rack of containers contains at least 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48, or 96 containers; A system or device according to any one of claims 1 to 4.

7. the contacting means is a pipette having a lumen, the system or device being configured to aspirate a liquid sample for analysis into the lumen, and the one or more electrochemical aptamer-based sensors being disposed within the lumen. A system or device according to any one of claims 1 to 6.

8. each of the plurality of separate liquid samples for analysis has its own dedicated pipette; 8. A system or apparatus according to claim 7.

9. the pipette comprising: (i) one or more electrical conduits for connecting the power source to the one or more electrochemical aptamer-based sensors; and (ii) one or more electrical conduits configured to carry an output current from the one or more electrochemical aptamer-based sensors.

9. A system or apparatus according to claim 7 or 8.

10. a pipette mount, the pipette being attachable to and detachable from the pipette mount; A system or device according to any one of claims 7 to 9.

11. the pipette mount includes an electrical connector configured to connect to (i) the one or more electrical conduits of the pipette for connecting the power source to the one or more electrochemical aptamer-based sensors, and (ii) the one or more electrical conduits configured to carry an output current from the one or more electrochemical aptamer-based sensors; 11. A system or apparatus according to claim 10.

12. a liquid aspirating means configured to aspirate a liquid sample from outside the pipette into the lumen of the pipette; A system or device according to any one of claims 7 to 11.

13. the contacting means is the sample holding means, and the one or more electrochemical aptamer-based sensors are disposed within or around the sample holding means so as to contact a liquid sample held therein; A system or device according to any one of claims 1 to 6.

14. the sample holding means comprising: (i) one or more electrical conduits for connecting the power source to the one or more electrochemical aptamer-based sensors; and (ii) one or more electrical conduits configured to carry an output current from the one or more electrochemical aptamer-based sensors.

14. A system or apparatus according to claim 13.

15. a sample holder mount, the sample holder being attachable to and detachable from the sample holder mount; 15. A system or apparatus according to claim 13 or 14.

16. the sample holder mount includes an electrical connector configured to connect to (i) the one or more electrical conduits of the sample holder for connecting the power source to the one or more electrochemical aptamer-based sensors, and (ii) the one or more electrical conduits configured to carry an output current from the one or more electrochemical aptamer-based sensors; 16. A system or apparatus according to claim 15.

17. each of said plurality of distinct liquid samples for analysis is contacted with a dedicated electrochemical aptamer-based sensor; 17. A system or apparatus according to any one of claims 13 to 16.

18. the one or more electrochemical aptamer-based sensors are in the form of one or more biosensor probes contactable with each of the plurality of distinct liquid samples for analysis; 18. A system or apparatus according to any one of claims 1 to 17.

19. the contacting means is a transferring means configured to bring together the one or more biosensor probes and the plurality of separate liquid samples for analysis; 20. A system or apparatus according to claim 18.

20. the moving means includes a motorized mechanism configured to lower the one or more biosensor probes toward the holding means into contact with the plurality of separate liquid samples for analysis, and / or to raise the holding means toward the one or more biosensor probes into contact with the plurality of separate liquid samples for analysis.

20. A system or apparatus according to claim 19.

21. each of said plurality of distinct liquid samples for analysis is contacted with a dedicated electrochemical aptamer-based sensor; 21. A system or apparatus according to any one of claims 18 to 20.

22. a processor configured to receive current values ​​or derivatives thereof output by the one or more electrochemical aptamer-based sensors and convert the current values ​​or derivatives thereof into clinically relevant values.

22. A system or apparatus according to any one of claims 1 to 21.

23. the clinically relevant value is the amount or concentration of the target analyte; 23. A system or apparatus according to claim 22.

24. and program instructions stored in an electronic memory, the program instructions configured to convert the current value or a derivative thereof into a clinically relevant value.

24. A system according to claim 22 or claim 23.

25. a networking module in operative communication with the processor, the networking module in network communication with a server in an analytical laboratory, the networking module transmitting the current value or its derivative value, or the clinically relevant value to the server in association with a sample identifier; 25. A system according to any one of claims 22 to 24.

26. configured as a sample processing and analysis robot; 26. A system or apparatus according to any one of claims 1 to 25.

27. is a substantially self-contained device, 27. A system or apparatus according to any one of claims 1 to 26.

28. the substantially self-contained device being sized to be suitable for placement on a laboratory bench; 28. A system or apparatus according to claim 27.

29. 1. The collocation of a pipette, a sample vessel, or a probe, each of said pipette, sample vessel, or probe including one or more electrochemical aptamer-based sensors configured to detect a target analyte.

30. the pipettes, sample vessels or probes are spatially arranged in an orderly manner so as to be usable by a robotic sample processing and analysis system or device; 30. The collocation of claim 29.

31. The regular arrangement is a regular spacing between the pipettes, sample containers or probes.

31. The collocation of claim 30.

32. The regular arrangement is an array arrangement or a grid arrangement.

32. The collocation of claim 30 or 31.

33. 33. A co-location device according to any one of claims 29 to 32, configured to be operable with a system or device according to any one of claims 1 to 28.

34. 1. A method for simultaneously analyzing multiple samples for one or more target analytes, the method comprising: placing each of a plurality of separate liquid samples for analysis in a separate container; contacting each of the separate liquid samples for analysis with one or more electrochemical aptamer-based sensors; receiving a current value output from each of the one or more electrochemical aptamer-based sensors; converting the current value or its derivative into a clinically relevant value; A method comprising:

35. At least about 2, 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 36, 48, or 96 samples are analyzed simultaneously; 35. The method of claim 34.

36. the one or more electrochemical aptamer-based sensors are disposed in or around a pipette tip configured to aspirate a liquid sample, or in or around a probe configured to contact a liquid sample, or in or around a container configured to hold a liquid sample; 36. The method of claim 34 or 35.

37. 37. A method according to any one of claims 34 to 36, at least in part performed by a system or device according to any one of claims 1 to 28.

38. 38. A method according to any one of claims 34 to 37, using a collocation according to any one of claims 26 to 30.

39. The method lacks or does not require any one or more of the following steps: sample preparation, dilution, centrifugation, purification, reagent addition, washing, mixing, incubation, heating, cooling, light detection, light spectroscopy, light scattering analysis, light absorption analysis, visual inspection, elution, radiation detection, inter-container sample transfer, and inter-device sample transfer; 39. The method of any one of claims 34 to 38.

40. 1. A method for assembling a customized set of pipettes, sample containers or probes capable of detecting one or more analytes in a sample, the method comprising: providing a library comprising a plurality of pipettes, sample vessels or probes, each of said pipettes, sample vessels or probes having an associated electrochemical aptamer-based sensor capable of detecting a particular analyte; selecting two or more pipettes, sample vessels or probes from an associated library; juxtaposing the two or more selected pipettes, sample containers or probes; A method comprising:

41. 1. A method of manufacturing a customized plate comprising a plurality of wells, each well capable of detecting one or more analytes in a sample, the method comprising: providing a library comprising a plurality of electrochemical aptamer-based sensors capable of detecting a particular analyte; selecting two or more of the electrochemical aptamer-based sensors; associating each of the wells with one of the selected electrochemical aptamer-based sensors; A method comprising:

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