WO2021226145A1 - Methods and devices for pathogen detection - Google Patents

Abstract

This invention relates to methods and devices for detecting the presence of pathogens in samples, particularly to detecting the presence of pathogen or antigens in samples, and more particularly to detecting the presence of pathogen or antigens in samples by detection of a binding event between a functional ligand, such as an aptamer, to a pathogen or antigen in a sample. Some exemplary embodiments of the invention include a kit with the use aptamers for an antigen, such as the spike (S) protein of the SARS-CoV-2 virus, in conjunction with an electrochemical sensor in a detection module that interfaces with a sample chamber.

Description

METHODS AND DEVICES FOR PATHOGEN DETECTION

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit and priority of U.S. provisional patent application Ser. No. 63/019,760, filed May 4, 2020, entitled “METHODS AND DEVICES FOR PATHOGEN DETECTION”, the contents of which are hereby incorporated by reference in its entirety.

FIELD OF THE INVENTION

[0002] This invention relates to methods and devices for detecting the presence of pathogens in samples, particularly to detecting the presence of pathogen or antigens in samples, and more particularly to detecting the presence of pathogen or antigens in samples by detection of a binding event between a functional ligand, such as an aptamer, to a pathogen or antigen in a sample.

BACKGROUND OF THE INVENTION

[0003] Aptamers, which are nucleic acid ligands capable of binding to molecular targets, have recently attracted increased attention for their potential application in many areas of biology and biotechnology. They may be used as sensors, therapeutic tools, to regulate cellular processes, as well as to guide drugs to their specific cellular target(s). Contrary to the actual genetic material, their specificity and characteristics are not directly determined by their primary sequence, but instead by their secondary and/or tertiary structure. Aptamers have been recently investigated as immobilized capture elements in a microarray format. Others have recently selected aptamers against whole cells and complex biological mixtures. Aptamers are typically characterized by binding to their target molecules via non-Watson-Crick (i.e. non-hybridization) mechanisms, such as by intermolecular forces resulting from the secondary or tertiary structure of the aptamer. This is especially true of non- nucleic acid target molecules where Watson-Crick mechanisms typically do not apply.

[0004] Aptamers are commonly identified by an in vitro method of selection sometimes referred to as Systematic Evolution of Ligands by Exponential enrichment or “SELEX’. SELEX typically begins with a very large pool of randomized polynucleotides which is generally narrowed to one aptamer ligand per molecular target. Once multiple rounds (typically 10-15) of SELEX are completed, the nucleic acid sequences are identified by conventional cloning and sequencing. Aptamers have most famously been developed as ligands to important proteins, rivaling antibodies in both affinity and specificity, and the first aptamer-based therapeutics are now emerging. More recently, however, aptamers have been also developed to bind small organic molecules and cellular toxins, viruses, and even targets as small as heavy metal ions. After identification of an aptamer sequence from sequencing after SELEX, the aptamer is typically manufactured afterwards in manners utilized with any other oligonucleotide, such as by standard synthesis methods, such as standard commercial nucleic acid synthe^⁰, eoti de synthesis by phosphoramidite method, etc. ), 1/030734 , _ay be utilized with biological synthesis methods, such polymerase chain reaction (PCR) or the like.

[0005] Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The disease was first identified in December 2019 in Wuhan, China, and has since spread globally, resulting in a coronavirus pandemic. Common symptoms include fever, cough, and shortness of breath. Other symptoms may include fatigue, muscle pain, diarrhea, sore throat, loss of smell, and abdominal pain. The time from exposure to onset of symptoms is typically around five days but may range from two to fourteen days. While the majority of cases result in mild symptoms, some patients progress to viral pneumonia and multi-organ failure. The virus is primarily spread between people during close contact, often via small droplets produced by coughing, sneezing, or talking. While these droplets are produced when breathing out, they usually fall to the ground or onto surfaces rather than being infectious over long distances. People may also become infected by touching a contaminated surface and then touching their eyes, nose, or mouth. The virus can survive on surfaces up to 72 hours. It is most contagious during the first three days after the onset of symptoms, although spread may be possible before symptoms appear and in later stages of the disease. The role of these asymptomatic carriers in transmission is not yet fully known; however, preliminary evidence suggests that they may contribute to the spread of the disease.

[0006] The standard method of diagnosis is by real-time reverse transcription polymerase chain reaction (rRT-PCR) from a nasopharyngeal swab. Chest CT imaging may also be helpful for diagnosis in individuals where there is a high suspicion of infection based on symptoms and risk factors; however, it is not recommended for routine screening. Current testing for the presence of the virus are experiencing issues, such as due to scale of sample collection, personnel required to perform the task, centralized lab processing with reporting of results subjected to extended delays, etc. Additionally, the availability of tests and lab capacity are often limiting availability to only those experiencing symptoms, far after they have likely been contagious. Current more rapid testing methods include detection of antibodies which the body produces in response to viral infections (e.g. IgG, IgM), but antibodies may only be typically present in detectable levels days or weeks after infection and may thus result in false negative results of a person being infected and having limited usefulness in early stage detection.

SUMMARY OF THE INVENTION

[0007] This invention relates to methods and devices for detecting the presence of pathogens in samples, particularly to detecting the presence of pathogen or antigens in samples, and more particularly to detecting the presence of pathogen or antigens in samples by detection of a binding event between a functional ligand, such as an aptamer, to a pathogen or antigen in a sample.

[0008] In general, a method and device for detecting the presence of a pathogen in a sample may include collecting a sample using a sampling component, placing the sample in a sample chamber and contacffi§ 2021/226145 a detector module containing a functional ligand thEP /U^P ! P A^iensor. The sample may generally include sputum, saliva, nasal mucus, or other fluid that may contain a pathogen, such as a virus (e.g. SARS-CoV-2, SARS-CoV-1, MERS-CoV, Ebola, influenza, etc.). The sample may be collected by use of a sampling component, such as a nasal swab or similar implement, directly deposited into a sample chamber, or a combination. The sample chamber may then be interfaced with the detector module, such as by insertion or other form of interface, such that the sample may contact a functional ligand that may bind to a pathogen or an antigen of the pathogen and cause a detectable binding event through interaction with a sensor to produce a signal. The signal may then be utilized to determine the presence and/or abundance of the pathogen or antigen in the sample. [0009] In one aspect of the invention, a device for detecting the presence of a pathogen in a sample may utilize a detector module having a body with a sample chamber interface, at least one functional ligand, such as an aptamer, that interacts with a sensor within said body, and a detection indicator for displaying or alerting a user of a positive or negative detection event, and/or the abundance of a detected pathogen or antigen.

[0010] In some exemplary embodiments, the functional ligand may include at least one nucleic acid aptamer with selective binding affinity to a pathogen or an antigen of a pathogen, such as, for example, the spike (S), envelope (E), and/or membrane (M) proteins of the SARS-CoV-2 virion, and/or to the whole virion itself.

[0011] In some exemplary embodiments, the functional ligand may also include a functional group (e.g. a redox reporter such as methylene blue (MB)) and be coupled to an electrochemical sensor such that binding of the antigen or pathogen to the functional ligand causes a conformational change that alters the orientation and/or distance of the functional group relative to the electrochemical sensor to generate a detectable signal, such as a change in the current measured from an electrode due to a change in electron transfer from a functional group to the electrode.

[0012] In some embodiments, signal enhancing features may be utilized to boost the signal from the binding of the antigen or pathogen to the functional ligand.

[0013] In another aspect of the invention, the components of the device may be provided as a kit and at least a portion of the components may be directly disposable after a single use such that the incidence of false results due to reuse or cross-contamination may be reduced or eliminated. The device also provides for a minimal number of components and may include a design for minimal or no transfer of sample after depositing into a sample chamber, which may be desirable to reduce the risk of transfer- related errors and/or contamination risk.

[0014] In some embodiments, the detector module may be disposable. In other embodiments, the detector module may be at least partially reusable. For example, the detector module may be constructed to be sterilizable and/or provide for appropriate replacement parts, such as, for example, the sensor. [0015]^wo ?!!²]/22<Vl -,ibodi merits, the detector module may also include a such that results may be transmitted, such as to a computer, mobile device or cloud service.

[0016] In general, a method for generating functional biomolecules includes obtaining a library, such as a diverse or randomized library, for example, of biomolecules. Biomolecules may generally include nucleic acids, particularly single-stranded nucleic acids, peptides, other biopolymers and/or combinations or modifications thereof. A library of biomolecules may include nucleic acid sequences, such as ribonucleic acid (RNA), deoxyribonucleic acid (DNA), artificially modified nucleic acids, and/or combinations thereof. The method for generating functional biomolecules further includes contacting the library of biomolecules with more than one target, such as, for example, a molecular target, material and/or substance. In general, the members of the library that do not bind with some affinity to the more than one target may be washed or otherwise partitioned from the remainder of the library, which may have a given level of binding affinity to the more than one target. The process may be repeated to partition the strongest binding members of the library. Amplification of the biomolecules may also be utilized to increase the numbers of the binding members of the library for subsequent repetitions and for isolation and/or purification of any final products of the process. Embodiments of the SELEX method may generally be utilized to achieve the generation of functional biomolecules of a given binding affinity, such biomolecules generally referred to as aptamers or ligands.

[0017] The present invention together with the above and other advantages may best be understood from the following detailed description of the embodiments of the invention and as illustrated in the drawings. The following description, while indicating various embodiments of the invention and numerous specific details thereof, is given by way of illustration and not of limitation. Many substitutions, modifications, additions or rearrangements may be made within the scope of the invention, and the invention includes all such substitutions, modifications, additions or rearrangements.

BRIEF DESCRIPTION OF THE FTGTTRES

[0018] FIG. 1 illustrates an embodiment of a kit containing a sampling component, a sample chamber and a detector module of the present invention;

[0019] FIGs. la, lb, lc, Id and le illustrate the use and operation of the kit of FIG. 1 to detect the presence of a pathogen in a sample;

[0020] FIG. 2 illustrates the principle of operation of a functional ligand binding to a target to generate a signal from an electrochemical sensor;

[0021] FIGs. 2a and 2b illustrate the use of additional functional ligands to enhance signal from binding of the target; and

[0022] FIG. 3 illustrates an exploded view of a detector module of the present invention.

[0023] The detailed description set forth below is intended as a description of the presently exemplified systems, devices and methods provided in accordance with aspects of the present invention and are not intended to represent the only forms in which the present invention may be prepared or utilized. It is to be understood, rather, that the same or equivalent functions and components may be accomplished by different embodiments that are also intended to be encompassed within the spirit and scope of the invention. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this invention belongs. Although any methods, devices and materials similar or equivalent to those described herein can be used in the practice or testing of the invention, the exemplary methods, devices and materials are now described. All publications mentioned herein are incorporated herein by reference for the purpose of describing and disclosing, for example, the designs and methodologies that are described in the publications which might be used in connection with the presently described invention. The publications listed or discussed above, below and throughout the text are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the inventors are not entitled to antedate such disclosure by virtue of prior invention.

[0024] This invention relates to methods and devices for detecting the presence of pathogens in samples, particularly to detecting the presence of pathogen or antigens in samples, and more particularly to detecting the presence of pathogen or antigens in samples by detection of a binding event between a functional ligand, such as an aptamer, to a pathogen or antigen in a sample.

[0025] In general, a method and device for detecting the presence of a pathogen in a sample may include collecting a sample using a sampling component, placing the sample in a sample chamber and contacting the sample with a detector module containing a functional ligand that interacts with a sensor. FIG. 1 illustrates a kit including a sampling component 102, a sample chamber 110 and a detector module 120, which may be provided in a container or other packaging, such as illustrated with a bag 90.

[0026] In one aspect of the invention, a device for detecting the presence of a pathogen in a sample may utilize a detector module having a body with a sample chamber interface, at least one functional ligand, such as an aptamer, that interacts with a sensor within said body, and a detection indicator for displaying or alerting a user of a positive or negative detection event, and/or the abundance of a detected pathogen or antigen. The sample may generally include sputum, saliva, nasal mucus, or other fluid that may contain a pathogen, such as a virus (e.g. SARS-CoV-2, SARS-CoV-1, MERS-CoV, Ebola, influenza, etc.). The sample may be collected by use of a sampling component, such as a nasal swab or similar implement, to swab a tissue of a person to collect fluid, such as from the nasal cavity, mouth, throat, etc., as shown with the sampling component 102 illustrated as a nasal swab with an absorbent end 102a, directly deposited into a sample chamber 110, or a combination. As illustrated in FIGs. temple chamber 110 may generally include a vessel, asPPJ V^^ Q^QZ^iiollow cylindrical vessel 112, for holding a fluid sample which may be deposited through the opening 111 after removing the cap 114, as shown with the sampling component 102 being inserted in FIG. la. Saliva, sputum or other fluid may also be deposited directly into the opening 111, such as by spitting, expectorating or the like to provide the sample. The sampling component 102 may also be used to stir or otherwise transfer the sample into the fluid, which may, for example, be utilized to aid in concentrating any pathogen or antigens thereof in the sample chamber 110. The sample chamber base 116, as illustrated with a flared base in FIGs. 1 and la, may be utilized to provide a stable base for the sample chamber 110 during sample collection. The base 116 may also be hollow and provide an enlarged reservoir for the sample fluid.

[0027] The sample chamber 110 may then be interfaced with the detector module 120, such as by insertion or other form of interface, as illustrated with the detector module 120 being placed on the opening 111 after sample collection with the hollow cylindrical vessel 112 at least partially inserting into the interface opening 124 in the body 122 of the detector module 120, as shown in FIGs. 1 and lb. The detector module 120 may be activated prior to or after insertion of the sample chamber 110, such as by actuating a power control, as shown with power button 126 in FIGs. 1 and lb. The power button 126 may further include a light or other indicator to show that the power is on, such as using an LED or other light source, as shown with power light indicator 136 in FIG. 3. As the fluid sample may generally collect toward the bottom of the sample chamber 110 due to gravity (e.g. toward the base 116) and to aid in containing the sample, after placement of the detector module 120, the device 100 may be inverted to cause gravity to draw the fluid sample toward the interface opening 124 in direction A and into the detector module 120, as illustrated in FIG. lc. The sample may then contact a sensing apparatus in the detector module 120 to produce a signal which is interpreted into a result to be relayed to the user, such as, for example, a positive or negative signal, as illustrated with the positive result indicator 128 lighting in FIG. Id, or with the negative result indicator 127 lighting in FIG. le. In some embodiments, other signals may be utilized, such as digital displays, audio signals, output to a wired or wireless device and/or any other appropriate signaling.

[0028] The sensing apparatus in the detector module 120 may generally include a functional ligand that may bind to a pathogen or an antigen of the pathogen and cause a detectable binding event through interaction with a sensor to produce a signal. The sensing apparatus may utilize any appropriate sensing modality, such as, for example, electrochemical sensor, fluorescent assay, enzyme-linked aptamer sorbent assay (ELASA), gold nanoparticle assay, or any other appropriate modality or combination thereof. The signal may then be utilized to determine the presence and/or abundance of the pathogen or antigen in the sample. In some exemplary embodiments, the functional ligand may include at least one nucleic acid aptamer with selective binding affinity to an antigen of a pathogen, such as, for example, the spike (S), envelope (E), and/or membrane (M) proteins of the SARS-CoV-2 virion, and/or to the whole virion itself. [0029] ?Q¾/226145jtrates a schematic of a sensor 140 utilizing an embodi!^J P§?¾i/9:?£l¾imical detection. In some exemplary embodiments, a functional ligand, as illustrated with aptamer 148 may be attached to an electrode surface 142, as illustrated with a gold electrode. The functional ligand may be attached by any appropriate method, such as by direct attachment with a functional linkage or group such as a thiol or di-thiol group, or any other appropriate attachment. In some embodiments, a capture oligonucleotide 147 may be attached to the electrode surface 142, as illustrated in FIG. 2 via thiol group 147a. The capture oligonucleotide 147 may then be utilized to hybridize to a portion of an aptamer 148 to attach the aptamer to the electrode surface 142. This may be desirable as the hybridization may be reversible and if desired, many different aptamers 148 may be designed to hybridize to the capture oligonucleotide 147, such that multiple different potential pathogens may be detected using a standardized detector module 120 by hybridizing an appropriate aptamer (or if needed, removing an already hybridized aptamer and replacing it with a different one). In some embodiments, the sensor 140 may be provided with an amount of fluid to wet and hydrate the functional ligands prior to introduction of the sample, such as buffer, water or other appropriate fluid. A seal may be included on the detector module 120 to keep the fluid in the interface opening 124 before use. In other embodiments, the functional ligands may be provided without fluid, such as, for example, by lyophilizing the aptamer 148 and/or the capture oligonucleotide 147 after attachment. The functional ligand may also include a functional group (e.g. a redox reporter such as methylene blue (MB)) and be coupled to an electrochemical sensor such that binding of the antigen or pathogen to the functional ligand causes a conformational change that alters the orientation and/or distance of the functional group relative to the electrochemical sensor to generate a detectable signal, such as a change in the current measured from an electrode due to a change in electron transfer from a functional group to the electrode.

[0030] FIG. 2 illustrates the aptamer 148 with a functional group 148a (illustrated as methylene blue redox reporter), where with no target 80 present (i.e. the pathogen or antigen), the aptamer 148 is in a first conformation (illustrated as being relatively away from the electrode surface 142 causing slow electron transfer (i.e. low current) between the functional group I48a and the electrode surface 142). When target 80 is present, such as when the antigen or pathogen is introduced via the sample from the sample chamber 110, the target 80 may bind to the aptamer 148, which may, generally and without being bound to any particular theory, a conformational change to a second conformation that alters the orientation and/or distance of the functional group 148a to the electrode surface 142, causing a change in the current (e.g. faster electron transfer from the functional group 148a to the electrode surface 142 (i.e. higher current) due to increased proximity). In general, the current from the electrochemical sensor may be determined as a change relative to a baseline, such as by reading the current of the electrode surface 142 versus that read at a reference electrode 144 (e.g. a potentiostat, such as a reference silver/silver chloride redox reaction), which is illustrated in FIG. 3. [003 l]WO 2021/226145 ibodiments, signal enhancing features may be uti 1 i from the binding of the antigen or pathogen to the functional ligand. For example, additional functional ligands may be utilized, such as, for example, to enhance or boost the signal generated by the binding of the antigen or pathogen. FIGs. 2a and 2b illustrate an example of additional functional ligands (shown as secondary aptamers 150 that are not immobilized on the electrode surface 142 may be exposed to the sample. Because multiple functional ligands (e.g. secondary aptamers 150) may be designed or developed to sterically bind a single virion, as shown with the multiple secondary aptamers 150 binding the target 80 (shown as a virion), such an approach may be used for increased signal, such as by, for example, increasing the number of functional groups 152 per target 80. In some embodiments, additional redox reporters may be utilized as the functional groups 152, thus multiplying the current change per target 80.

[0032] In some embodiments, the functional groups 152 may be utilized in for enzyme-linked assays, such as with a biotin which may then bind to a biotin-binding moiety (e.g. streptavidin, avidin, etc.) linked to an enzyme for a reporter (e.g. horseradish peroxidase, etc.), or vice versa.

[0033] In some embodiments, additional functional ligands may be present on particles which may be utilized to draw down the target 80 to the surface of the sensor, such as illustrated with particles 154 with secondary aptamers 156. The particles 154 may, for example, be utilized to attach mass to the target 80 via binding by the secondary aptamers 156. The additional mass may be utilized to aid in settling of the target 80, such as by gravity or centrifugation. Secondary aptamers 150 with functional groups 152 may also be present to increase signal by binding the target 80 to form aggregates that are pulled down by the magnet.

[0034] In some embodiments, the particles 154 may include, for example, magnetic properties (e.g. magnetic nanoparticles) such that the application of magnetic force may be utilized to move the bound target 80, such as by placement of a magnet 149 proximal to the sensor to draw the target 80 via the particles 154 to a sensor (illustrated in FIG. 2b as electrode surface 142). Secondary aptamers 150 with functional groups 152 may also be present to increase signal by binding the target 80 to form aggregates that are pulled down by the magnet.

[0035] In some embodiments, a wash step may be incorporated to wash away unbound functional ligands, such as the secondary aptamers 150, such that background signal may be reduced. For example, after binding between the target 80 and the aptamers 148 on the sensor occurs, excess fluid containing unbound secondary aptamers 150 may be removed or segregated, such as by introducing a wash buffer and removing, by inverting the sample chamber 110 to gather the excess fluid (additionally the detector module 120 with the target 80 bound to the aptamers 148 may be removed from the sample chamber 110 and placed with another chamber with wash buffer, inverted again to wash, and/or reinverted for reading the signal). This may generally remove excess unbound secondary aptamers 150 such that the extra functional groups 152 may not enhance the signal. [0036]^WO module 120 may further include features to provideP£T/JS¾²9?A/PA97A⁴ent for the sample and sensor 140 to interact. For example, seals, gaskets or the like may be utilized to aid in sealing the hollow cylindrical vessel 112 against the sensor 140 to produce a fluid-tight chamber, such as using sealing members 124a, 124b. In embodiments where the functional ligands are provided dry prior to use, the fluid from the sample may then collect on the sensor 140 and may generally wet and hydrate the functional ligands for operation of the sensor 140.

[0037] In general, the sensor 140 and other operative components of the detector module 120 may be housed with the body 122, such as with the upper and lower halves 122a, 122b in FIG. 3. The sensor 140 may further include output contacts 146 that provide current to the sensor 140 and carry the output signal to a processing device, as shown with the leads 132 on the processing device 130 in FIG. 3. The processing device 130 may generally include a logic and/or memory module 134 to control the overall operation of the detection module 120, and a power source, as illustrated with batteries 135, to power the processing device 130 and to provide current to the sensor 140 via leads 132. In some embodiments, to accommodate the components within the body 122 and to enable the sample to pass through and reach the sensor 140, the processing device 130 may include an aperture 124c to allow the sample to pass through. Upon introduction of the sample to the sensor 140, the processing device 130 may generally detect a change in signal due to presence of the target 80 (i.e. the pathogen or antigen), or no change relative to the baseline due to the target 80 not being present or present below a detection threshold. The processing device 130 may then output the result to the user, such as with the illustrated light sources 137, 138 (e.g. LEDs) to illuminate the negative or positive result indicators 127, 128, respectively.

[0038] In another aspect of the invention, the components of the device 100 may be provided as a kit and at least a portion of the components may be directly disposable after a single use such that the incidence of false results due to reuse or cross-contamination may be reduced or eliminated. For example, the device 100 may be provided in a sealed container or packaging, such as the bag 90 in FIG. 1 and after use, the device 100 (all components and the sample), may be disposed of, such as by replacing in the bag 90 which may be resealed or closed to aid in containment of potential contaminants and then safely disposed. This may also generally reduce the cost from using reusable components that may require sterilization or other actions to prepare for further uses. The device also provides for a minimal number of components and may include a design for minimal or no transfer of sample after depositing into a sample chamber, which may be desirable to reduce the risk of transfer-related errors and/or contamination risk. For example, as illustrated in FIGs. la, lb and lc, after the sample is deposited in the sample chamber 110, the detector module 120 is placed on it and the sample does not move to another container.

[0039] In some embodiments, the detector module 120 may be disposable. In other embodiments, the detector module 120 may be at least partially reusable. For example, the detector module 120 may be con$fQ 2¾V.?!⁶J3? rilizable and/or provide for appropriate replacement paP£T/U¾9¾ 9 P ¾mple, the sensor 140 being replaced or washed to remove any residual sample.

[0040] In some embodiments, the detector module 120 may also include a communication link such that results may be transmitted, such as to a computer, mobile device or cloud service. The communication link may utilize a wired or wireless connection, such as WiFi or LTE. The transmitted results may be desirable, for example, in generating mapping of results and/or for geographical tracking.

[0041] Although the invention has been described with respect to specific embodiments thereof, these embodiments are merely illustrative, and not restrictive of the invention. The description herein of illustrated embodiments of the invention, including the description in the Abstract and Summary, is not intended to be exhaustive or to limit the invention to the precise forms disclosed herein. Rather, the description is intended to describe illustrative embodiments, features and functions in order to provide a person of ordinary skill in the art context to understand the invention without limiting the invention to any particularly described embodiment, feature or function, including any such embodiment feature or function described in the Abstract or Summary. While specific embodiments of, and examples for, the invention are described herein for illustrative purposes only, various equivalent modifications are possible within the spirit and scope of the invention, as those skilled in the relevant art will recognize and appreciate. As indicated, these modifications may be made to the invention in light of the foregoing description of illustrated embodiments of the invention and are to be included within the spirit and scope of the invention. Thus, while the invention has been described herein with reference to particular embodiments thereof, a latitude of modification, various changes and substitutions are intended in the foregoing disclosures, and it will be appreciated that in some instances some features of embodiments of the invention will be employed without a corresponding use of other features without departing from the scope and spirit of the invention as set forth. Therefore, many modifications may be made to adapt a particular situation or material to the essential scope and spirit of the invention.

[0042] Reference throughout this specification to "one embodiment", "an embodiment", or "a specific embodiment" or similar terminology means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment and may not necessarily be present in all embodiments. Thus, respective appearances of the phrases "in one embodiment", "in an embodiment", or "in a specific embodiment" or similar terminology in various places throughout this specification are not necessarily referring to the same embodiment. Furthermore, the particular features, structures, or characteristics of any particular embodiment may be combined in any suitable manner with one or more other embodiments. It is to be understood that other variations and modifications of the embodiments described and illustrated herein are possible in light of the teachings herein and are to be considered as part of the spirit and scope of the invention. [0043]^WO ?ft²1/226145;ripti on herein, numerous specific details are p r o v i d T/ IIS 2 < ¹2 j /(*? 07 f, 1 e s of components and/or methods, to provide a thorough understanding of embodiments of the invention. One skilled in the relevant art will recognize, however, that an embodiment may be able to be practiced without one or more of the specific details, or with other apparatus, systems, assemblies, methods, components, materials, parts, and/or the like. In other instances, well-known structures, components, systems, materials, or operations are not specifically shown or described in detail to avoid obscuring aspects of embodiments of the invention. While the invention may be illustrated by using a particular embodiment, this is not and does not limit the invention to any particular embodiment and a person of ordinary skill in the art will recognize that additional embodiments are readily understandable and are a part of this invention.

[0044] As used herein, the terms “comprises,” “comprising,” "includes," "including," "has," "having," or any other variation thereof, are intended to cover a non-exclusive inclusion. For example, a process, product, article, or apparatus that comprises a list of elements is not necessarily limited only those elements but may include other elements not expressly listed or inherent to such process, process, article, or apparatus.

Claims

CLAIlWO 2021/226145 PCT/US2021/030734

1. A device for detecting the presence of a pathogen in a sample comprising: a detection module having a body with an interface aperture; a sensor connected to a fluid space connected to said interface aperture, said sensor comprising at least one aptamer having a specific binding affinity to at least a portion of a pathogen attached to an electrode; a processing device housed within said body and functionally connected to said sensor; and at least one output indicator adapted to relay a positive or negative signal to a user corresponding to the presence or non-presence of said pathogen, respectively.

2. The device of claim 1, further comprising a sample chamber having a fluid holding vessel that interfaces with said interface aperture.

3. The device of claim 2, further comprising a sampling component adapted for placement in said fluid holding vessel.

4. The device of claim 1, wherein said at least one aptamer further comprises a redox reporter functional group attached to an end of said aptamer.

5. The device of claim 1, wherein said at least one aptamer is attached to said electrode by hybridization to a capture oligonucleotide attached to said electrode.

6. The device of claim 1, further comprising a reference electrode on said sensor.

7. The device of claim 1, where said at least one aptamer binds to the spike (S) protein of the SARS-CoV-2 virus.

8. The device of claim 1, further comprising a plurality of secondary aptamers having a binding affinity to at least a portion of a pathogen and not attached to said sensor.

9. The device of claim 8, wherein said secondary aptamers comprise a functional group selected from the group consisting of a redox reporter functional group, biotin and a biotin-binding moiety.

10. The device of claim 8, further comprising a plurality of particles comprising additional said secondary aptamers.

11_. claim 10, further comprising a magnet disposed pnPCT/US202lM¾p4_ensor wherein said particles comprise a magnetic nanoparticle.

12. A method for detecting the presence of a pathogen in an organism comprising: collecting a fluid sample potentially containing a pathogen from an organism; depositing said fluid sample in a sample chamber having an open top end and a closed base; interfacing said sample chamber with a detector module comprising: a body with an interface aperture; a sensor connected to a fluid space connected to said interface aperture, said sensor comprising at least one aptamer having a specific binding affinity to at least a portion of a pathogen attached to an electrode; a processing device housed within said body and functionally connected to said sensor; and at least one output indicator adapted to relay a positive or negative signal to a user corresponding to the presence or non-presence of said pathogen, respectively; contacting said fluid sample with said sensor in said fluid space; wherein said sensor generates a signal response to the binding of at least a portion of said pathogen to said at least one aptamer and said signal response causes said processing device to output a positive signal to said user.

13. The method of claim 12, wherein said detector module interfaces with said sample chamber by placement of said interface aperture over said open top end and inverting said sample chamber and detector module to cause said fluid sample to move toward said open top end and into said detector module.