Multiplied sensor device and method for manufacturing the same

Abstract

Provided are an electrochemical sensor device that introduces a needle electrode into the skin of a subject and contacts the body fluid or tissue of the subject to detect a target analyte, and a method for manufacturing the device. This device has a movable part that pushes a needle electrode into the skin of a subject, at least two needle electrodes are selected from a library, and the device is provided with multiplexing ability.

Description

Technical Field

[0001] The present invention generally relates to sensor devices used for detecting test samples or analytes in vivo. In particular, the present invention relates to sensor devices manufactured by assembling a plurality of individual electrodes. The electrodes can be selected to include two or more working electrodes, and each working electrode can specifically detect different analytes or the same analyte.

Background Art

[0002] Electrochemical aptamer-based (EAB) sensors open the possibility of in situ and real-time monitoring of target analytes in a subject. In that context, an EAB sensor can include a needle-shaped working electrode coated with a redox-modified aptamer that can specifically bind to a target analyte. The working electrode can be inserted into the subject's skin to allow the aptamer to contact a biological fluid such as interstitial fluid or blood. An inspection potential is applied to the working electrode (e.g., by square wave voltammetry), and the current flowing through the electrode is measured. The amount of the measured current is used to determine the amount of the analyte present in the biological fluid.

[0003] In a clinical scenario, it may be desirable to simultaneously detect multiple target analytes in a subject's biological fluid. The amounts of multiple clinically relevant analytes in a particular disease or condition can be determined and combined to provide detailed diagnostic or prognostic information about the subject. For example, a subject suspected of having a myocardial infarction can have their blood tested for several endogenous cardiac markers such as troponin, creatine phosphokinase, C-reactive protein, myoglobin, etc.

[0004] In some cases, biological fluids may be analyzed for multiple drugs, etc., and multiple exogenous analytes in a subject undergoing combination therapy. In other situations, both endogenous and exogenous analytes may be detected simultaneously. One such situation is when a nephrotoxic antibiotic is administered and the amount of the drug and endogenous toxicity markers (such as liver enzymes) are analyzed.

[0005] In applications for detecting multiple analytes, it is necessary to prepare multiple sensor devices, which leads to the problem of increased cost. In the previous example, a first sensor device for detecting the drug and a second sensor device for detecting liver enzymes are required, and each device has a dedicated housing, electronic equipment, and power source. Furthermore, it is necessary to attach multiple sensor devices to the subject's body, which is time-consuming and inconvenient. Each sensor to be attached may also become a target for catching on the clothes and other objects in the subject's environment.

[0006] In the case of sensor technologies capable of sensing multiple analytes, each electrode is either independently functionalized or appropriately prepared in situ, resulting in a significant increase in manufacturing cost, time, and complexity.

[0007] As discussed above, the EAB sensor can have electrodes in the form of needles or microneedles. A single microneedle typically has a length of 150 - 1500 μm, a width of 50 - 250 μm, and a tip thickness of the tapered tip of 1 - 25 μm. The microneedle can be manufactured from metal, silicon, polymer, glass, or ceramic, and the base of the microneedle is typically attached to a base substrate to form an array. The base substrate of the microneedle may contain an adhesive to improve its engagement with the skin.

[0008] In the prior art, numerous devices for inserting microneedles into the skin of a subject have been disclosed. Such devices are typically configured so that a subject can apply the microneedles in a non-clinical environment such as at home. Ease of use and reproducibility are the main purposes of these devices.

[0009] Some devices are specialized only for the application of microneedles, and when the operation is completed, the device is removed together with the microneedles. Other prior art applicator devices are configured to be separated from the microneedles, so that the microneedles can remain in the skin for a certain period after introduction.

[0010] Yet another type of prior art device is configured to introduce microneedles, and the device (including the microneedles) remains in the original position of the subject for a certain period. These devices provide ease of use for the subject, but there are still some problems.

[0011] One problem is that such devices are generally conspicuous and are noticed by the subject immediately. The device may get caught on clothing or other objects nearby and may come off completely or partially. These devices need to be worn at night, and it causes great discomfort when the subject rolls over the device.

[0012] Furthermore, there is also the problem that prior art devices have a large number of individual parts and are complex. This increases the cost and the tendency to fail. The increase in the number of parts also increases the weight, which makes it more conspicuous to the subject. The discomfort related to the weight of the device has been found to increase in proportion to the wearing period. In some applications (such as hormone monitoring), continuous real-time data may be required for several weeks. During that period, the device may need to be replaced several times, but the problem of the subject wearing a heavy item for a long time remains.

[0013] Furthermore, there is also a problem that the subject may not be able to be sure whether the microneedle has been properly inserted into the skin initially, or even whether it has been properly embedded in the skin over time. Conventional sensor devices typically include a housing, the lower surface of which is flush with the surface of the skin. Due to the presence of the housing, it is difficult, if not impossible, for the subject to check whether the microneedle has been properly embedded by looking at the surface of the skin. In case of doubt, the device can be removed and a new device can be attached. When the microneedle has actually been properly inserted, the replacement is wasteful.

[0014] One aspect of the present invention is to provide an improvement of a conventional sensor device and a method for manufacturing the same. A further aspect of the present invention is to provide a useful alternative to a conventional sensor device and a conventional manufacturing method.

[0015] Discussions of documents, acts, materials, devices, articles, etc. are included in this specification only for the purpose of providing context for the present invention. None of these matters, either individually or in combination, are intended to suggest or indicate that any of them formed part of the basis of the prior art or were common general knowledge in the field related to the present invention that existed before the priority date of each claim of this application.

Summary of the Invention

[0016] In a first aspect, although not necessarily the broadest aspect, the present invention provides a method for manufacturing an electrochemical aptamer-based sensor device, the method including assembling two or more electrodes of an electrochemical aptamer-based sensor with a device for bringing the two or more electrodes into contact with the skin of a subject.

[0017] In an embodiment of the first aspect, at least one of the two or more electrodes is a working electrode configured to specifically detect an analyte.

[0018] In one embodiment of the first aspect, at least two of the two or more electrodes are each a working electrode configured to specifically detect an analyte.

[0019] In one embodiment of the first aspect, each of the two or more working electrodes contains a different aptamer species, and each of the different aptamer species is configured to specifically detect a different analyte or the same analyte.

[0020] In one embodiment of the first aspect, the two or more working electrodes are assembled with a fixed mutual spacing relationship.

[0021] In one embodiment of the first aspect, the method includes the step of assembling one or more counter electrodes with the two or more working electrodes.

[0022] In one embodiment of the first aspect, the method includes the step of assembling one or more reference electrodes with the two or more working electrodes.

[0023] In one embodiment of the first aspect, the two or more working electrodes and the electrodes are regularly arranged.

[0024] In one embodiment of the first aspect, the regular arrangement is an array.

[0025] In one embodiment of the first aspect, each of the two or more working electrodes is substantially equidistant from one of the one or more counter electrodes.

[0026] In one embodiment of the first aspect, each of the two or more working electrodes is substantially equidistant from one of the one or more reference electrodes.

[0027] In one embodiment of the first aspect, all the electrodes are arranged with a fixed mutual spatial relationship.

[0028] In one embodiment of the first aspect, the distance or average distance between the electrodes is less than about 10 millimeters (mm), 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, 2 mm, 1 mm, 0.9 mm, 0.8 mm, 0.7 mm, 0.6 mm, 0.5 mm, 0.4 mm, 0.3 mm, 0.2 mm, or 0.1 mm.

[0029] In one embodiment of the first aspect, all the electrodes are about 100 mm 2 , 90 mm 2 , 80 mm 2 , 70 mm 2 , 60 mm 2 , 50 mm 2 , 40 mm 2 , 30 mm 2 , 20 mm 2 , 10 mm 2 , 9 mm 2 , 8 mm 2 , 7 mm 2 , 6 mm 2 , 5 mm 2 , 4 mm 2 , 3 mm 2 , 2 mm 2 , or 1 mm 2 and are arranged within a region less than that.

[0030] In one embodiment of the first aspect, at least one of the two or more working electrodes, and / or at least one of the one or more counter electrodes, and / or at least one of the one or more reference electrodes is a wire, a needle, or a micro-needle.

[0031] In one embodiment of the first aspect, the assembly includes the step of attaching each electrode to a mounting portion.

[0032] In one embodiment of the first aspect, the mounting portion is substantially resistant to bending and / or stretching and / or contraction.

[0033] In one embodiment of the first aspect, the mounting portion electrically insulates each electrode from each other electrode.

[0034] In one embodiment of the first aspect, the electrode and / or the mount portion are configured to form a waterproof seal at the joint formed therebetween.

[0035] In one embodiment of the first aspect, the waterproof seal is formed by press-fitting, snap-fitting, or friction fitting between the electrode and the mount portion.

[0036] In one embodiment of the first aspect, the waterproof seal is formed by a flexible seal or a curable sealant applied to the joint or around it.

[0037] In one embodiment of the first aspect, the waterproof seal is formed by a screw connection between the electrode and the mount portion.

[0038] In one embodiment of the first aspect, at least one electrode includes an extended region configured to contact the surface of the mount portion.

[0039] In one embodiment of the first aspect, the method includes assembling at least 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 working electrodes.

[0040] In one embodiment of the first aspect, each of the electrodes is a wire, a needle, or a micro-needle.

[0041] In one embodiment of the first aspect, each of the working electrodes is obtained by removing a working electrode from a group of working electrodes having the same analyte specificity.

[0042] In one embodiment of the first aspect, the group of working electrodes is held in a holder configured to removably hold the electrodes.

[0043] In one embodiment of the first aspect, two or more working electrodes are selected from an electrode library including a plurality of working electrodes each containing a different aptamer species.

[0044] In one embodiment of the first aspect, electrodes containing the same aptamer species are grouped in individual holders or in regions of a single holder.

[0045] In one embodiment of the first aspect, a device for bringing two or more electrodes into contact with the skin of a subject includes a skin contact portion defining a skin contact surface and one or more spaces through which two or more electrodes can extend, and a movable portion configured to move two or more electrodes from a first position behind the skin contact surface to a second position protruding from the skin contact surface.

[0046] In one embodiment of the first aspect, the device includes a holding portion configured to keep the skin contact surface in contact with the skin during use.

[0047] In one embodiment of the first aspect, the movable portion is configured to move from the first position to the second position along a non-linear path.

[0048] In one embodiment of the first aspect, the non-linear path is a substantially arc-shaped path.

[0049] In one embodiment of the first aspect, the movable portion has a connection end and a free end.

[0050] In one embodiment of the first aspect, the free end moves a longer distance than the connection end.

[0051] In one embodiment of the first aspect, the non-linear path is drawn with reference to the free end.

[0052] In one embodiment of the first aspect, the non-linear path is less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, or 3 mm.

[0053] In one embodiment of the first aspect, the angle of the arc is less than about 45°, 40°, 35°, 30°, 25°, 20°, 15°, 14°, 13°, 12°, 11°, 10°, 9°, 8°, 7°, 6°, or 5°.

[0054] In one embodiment of the first aspect, the movable part has a swivel part, a hinge part, a bending part, or a mounting part.

[0055] In one embodiment of the first aspect, the movable part is associated with the mount part.

[0056] In one embodiment of the first aspect, during use, the mount part is fixed and the movable part is movable relative to the mount part.

[0057] In one embodiment of the first aspect, the mount part includes a portion that allows the movable part to swivel, hinge, bend, or attach.

[0058] In one embodiment of the first aspect, the mount part is disposed at a certain distance from the skin contact surface.

[0059] In one embodiment of the first aspect, the mount part is disposed at a distance of less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, or 2 mm from the skin contact surface.

[0060] In one embodiment of the first aspect, the mount part is substantially transverse to the movable part.

[0061] In one embodiment of the first aspect, the device further includes a release part operable by the user, the release part being configured to hold the movable part in a first position until the user actuates the release part, and when the user actuates the release part, the movable part is released and can move to a second position.

[0062] In one embodiment of the first aspect, the device further includes a locking part configured to lock the movable part when the movable part is in the second position.

[0063] In one embodiment of the first aspect, the device is configured such that a motive power generated inside and / or outside the device is required to move the movable part from the first position to the second position.

[0064] In one embodiment of the first aspect, the motive power inside the device is generated from a spring, an elastically deformable member, a shape memory member, or other biasing means, and the motive power outside the device is generated from a user.

[0065] In one embodiment of the first aspect, the device does not have an internal motive power generator configured to move the movable part from the first position to the second position.

[0066] In one embodiment of the first aspect, the holding part is a dermatologically acceptable composition disposed on or around the skin contact surface, or includes this composition.

[0067] In one embodiment of the first aspect, the dermatologically acceptable composition is an adhesive or its functional equivalent.

[0068] In one embodiment of the first aspect, the holding part is configured to mechanically hold the skin contact surface in contact with the skin.

[0069] In one embodiment of the first aspect, the holding part is selected from any one or more of a strap, a band, a belt, a clamp, a grip, a necktie, a fastener, a sleeve, a stocking, a sock, a glove, a cap, a hat, pants, a singlet, a shirt, a bra, a top, trousers, a scarf, a ring, glasses, and a choker.

[0070] In one embodiment of the first aspect, two or more electrodes are mechanically connected to the movable part directly or indirectly.

[0071] In one embodiment of the first aspect, two or more electrodes are wires, needles, and / or microneedles.

[0072] In one embodiment of the first aspect, two or more electrodes form an array.

[0073] In one embodiment of the first aspect, two or more electrodes are of a length sufficient to contact the epidermis, dermis, or subcutaneous tissue of the subject.

[0074] In one embodiment of the first aspect, two or more electrodes are configured to conduct an electric current to, from, or through the skin, conduct a sound wave to, from, or through the skin, conduct light to, from, or through the skin, conduct heat to, from, or through the skin, sample a body fluid or tissue from the skin, deliver a biologically active substance to the skin, or introduce an analyte sensing substance to the skin during use.

[0075] In one embodiment of the first aspect, two or more electrodes are each conductive, and the device further includes a circuit having an audio, visual, or tactile indicator, the circuit being configured to activate the indicator when one or more protrusions contact a conductive fluid naturally present on the skin.

[0076] In one embodiment of the first aspect, the circuit includes at least two protrusions, and the circuit is configured to be completed by activating the indicator when at least two protrusions contact a conductive fluid naturally present on the skin.

[0077] In one embodiment of the first aspect, the circuit includes one protrusion and at least one conductive pad disposed in contact with the skin, and the circuit is configured to be completed by electrically communicating the protrusion and the pad with a conductive fluid naturally present on the skin to activate the indicator.

[0078] In one embodiment of the first aspect, when the device is applied to the skin, the movable part is in the second position, and any part of each of two or more electrodes protruding from the skin contact surface is embedded in the skin, the housing extends at most about 5 mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 11 mm, 12 mm, 13 mm, 14 mm, 15 mm, 16 mm, 17 mm, 18 mm, 19 mm, or 20 mm or less over most or substantially all of the skin.

[0079] In one embodiment of the first aspect, the device is configured to be used over a period exceeding about 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 24 hours, 36 hours, 48 hours, 60 hours, 72 hours, 84 hours, or 96 hours.

[0080] In one embodiment of the first aspect, two or more electrodes are configured such that they cannot be separated from the device or cannot be separated without the assistance of a tool.

[0081] In one embodiment of the first aspect, the movable part and the mount part are integrated.

[0082] In one embodiment of the first aspect, the integrated movable part and mount part are manufactured from an elastically deformable material.

[0083] In one embodiment of the first aspect, the integrated movable part and mount part are part of the circuit board of the device.

[0084] In one embodiment of the first aspect, the movable part is biased towards the second position and is maintained in the first position against the bias by a user - operable release part until the release part is actuated. When the release part is actuated, the movable part is released and can move to the second position.

[0085] In one embodiment of the first aspect, the releasable portion operable by the user is a shelf configured to hold the movable portion in the first position, and the motive force provided by the user deforming the shelf and / or the movable portion enables the movable portion to be released from the shelf and move to the second position.

[0086] In one embodiment of the first aspect, the movable portion is hingedly connected in relation to the skin contact portion.

[0087] In one embodiment of the first aspect, the hinge is disposed in or towards the peripheral region of the movable portion and the skin contact portion.

[0088] In one embodiment of the first aspect, the releasable portion includes a member configured to maintain the movable portion in the first position, but is removable or deformable by the user such that the movable portion can move to the second position.

[0089] In one embodiment of the first aspect, the member is removable by sliding substantially across the skin contact portion.

[0090] In one embodiment of the first aspect, the member is substantially wedge-shaped, and the device has a hinge associating the movable portion with the skin contact portion, with the thin portion of the wedge disposed proximally to the hinge and the thick portion of the wedge disposed distally to the hinge.

[0091] In one embodiment of the first aspect, the releasable portion is removable from the device and includes a gripping portion to facilitate manual removal.

[0092] In the second aspect, the present invention provides an electrochemical aptamer-based sensor device including an assembly of two or more electrodes and a device for contacting the two or more electrodes with the skin of a subject.

[0093] In one embodiment of the second aspect, at least one of the two or more electrodes is a working electrode including an aptamer species configured to specifically detect an analyte.

[0094] In one embodiment of the second aspect, at least two of the two or more electrodes are working electrodes each containing a different aptamer species.

[0095] In one embodiment of the second aspect, the two or more electrodes are assembled with a fixed mutual spacing relationship.

[0096] In one embodiment of the second aspect, one of the two or more electrodes is a counter electrode.

[0097] In one embodiment of the second aspect, one of the two or more electrodes is a reference electrode.

[0098] In one embodiment of the second aspect, the electrodes are regularly arranged.

[0099] In one embodiment of the second aspect, the regular arrangement is an array.

[0100] In one embodiment of the second aspect, the two or more electrodes include a counter electrode and two or more working electrodes, and each of the two or more working electrodes is substantially equidistant from the counter electrode.

[0101] In one embodiment of the second aspect, the two or more electrodes include a reference electrode and two or more working electrodes, and each of the two or more working electrodes is substantially equidistant from the reference electrode.

[0102] In one embodiment of the second aspect, all the electrodes are arranged with a fixed mutual spatial relationship.

[0103] In one embodiment of the second aspect, the distance or average distance between the electrodes is less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, 2 mm, 1 mm, 0.9 mm, 0.8 mm, 0.7 mm, 0.6 mm, 0.5 mm, 0.4 mm, 0.3 mm, 0.2 mm, or 0.1 mm.

[0104] In one embodiment of the second aspect, all the electrodes are about 100 mm 2 、90 mm2 , 80 mm 2 , 70 mm 2 , 60 mm 2 , 50 mm 2 , 40 mm 2 , 30 mm 2 , 20 mm 2 , 10 mm 2 , 9 mm 2 , 8 mm 2 , 7 mm 2 , 6 mm 2 , 5 mm 2 , 4 mm 2 , 3 mm 2 , 2 mm 2 , or 1 mm 2 is disposed within a region less than.

[0105] In one embodiment of the second aspect, at least one of the two or more working electrodes and / or at least one of the one or more counter electrodes and / or at least one of the one or more reference electrodes is a wire, a needle, or a micro-needle.

[0106] In one embodiment of the second aspect, the electrode is attached to a mounting portion.

[0107] In one embodiment of the second aspect, the mounting portion is substantially resistant to bending and / or stretching and / or contraction.

[0108] In one embodiment of the second aspect, the mounting portion electrically insulates each electrode from each other electrode.

[0109] In one embodiment of the second aspect, the working electrode is formed separately from the mounting portion, and the working electrode and the mounting portion are assembled to form a device.

[0110] In one embodiment of the second aspect, the working electrode and / or the mounting portion is configured to form a waterproof seal at the joint formed therebetween.

[0111] In one embodiment of the second aspect, the waterproof seal is formed by press-fitting, snap-fitting, or friction fitting between the electrode and the mounting portion.

[0112] In one embodiment of the second aspect, the waterproof seal is formed by a flexible seal or a curable sealant applied to the joint or around it.

[0113] In one embodiment of the second aspect, the waterproof seal is formed by a screw connection between the electrode and the mounting portion.

[0114] In one embodiment of the second aspect, at least one electrode includes an extended region configured to contact the surface of the mounting portion.

[0115] In one embodiment of the second aspect, the device includes at least 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 working electrodes.

[0116] In one embodiment of the second aspect, each of the electrodes is a wire, a needle, or a micro-needle.

[0117] In one embodiment of the second aspect, each of the electrodes is obtained by removing working electrodes from a group of electrodes having the same aptamer species, dimensions, materials, or functions.

[0118] In one embodiment of the second aspect, the group of electrodes is held within a holder configured to removably hold the electrodes.

[0119] In one embodiment of the second aspect, at least one of the electrodes is a working electrode, and the working electrode is selected from an electrode library including at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 working electrodes each including a different aptamer species.

[0120] In one embodiment of the second aspect, working electrodes containing the same aptamer species are grouped in individual holders or in regions of a single holder.

[0121] In one embodiment of the second aspect, the device has the device characteristics defined in any embodiment of the first aspect that refers to the device.

[0122] In a third aspect, the present invention provides a system for manufacturing an electrochemical aptamer-based sensor device, the system including a library of two or more electrodes of the electrochemical aptamer-based sensor device and a mount configured to attach two or more working electrodes in a fixed mutual spacing relationship, the mount being provided by a device having the above characteristics.

[0123] In one embodiment of the third aspect, the two or more electrodes are each working electrodes, and each working electrode contains a different aptamer species.

[0124] In one embodiment of the third aspect, the electrode library includes at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 working electrodes containing different aptamer species.

[0125] In one embodiment of the third aspect, electrodes containing the same aptamer species specificity are grouped in individual holders or in regions of a single holder.

Brief Description of the Drawings

[0126]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6A

Figure 6B

Figure 7A

Figure 7B

Figure 8

Figure 9A

Figure 9B

Figure 10

Figure 11

Figure 12

Figure 13

Embodiments for Carrying out the Invention

[0127] After considering this description, those skilled in the art will appreciate how the present invention may be implemented in various alternative embodiments and alternative applications. However, while various embodiments of the present invention are described herein, it is understood that these embodiments are presented for purposes of illustration only and are not limiting. Accordingly, this description of various alternative embodiments should not be construed as limiting the scope or breadth of the present invention. Further, the description of advantages or other aspects is applicable to specific exemplary embodiments and not necessarily to all embodiments or any embodiments covered by the claims.

[0128] Throughout the description and claims of this specification, the words "comprise", "comprising", and variations such as "comprises" are not intended to exclude other additional elements, components, integers, or steps.

[0129] References throughout this specification to "one embodiment" or "an embodiment" mean that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, the appearances of the phrases "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.

[0130] As used herein, terms indicating position such as "lateral", "transverse", "above", "upper", "below", "higher", "lower", "upward", "downward", "plan view", etc. are considered in relation to a device in which the electrodes are oriented downward toward the ground.

[0131] As used herein, the article "a" or "an" refers to one or more (i.e., at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0132] As used herein, the terms "about" and "approximately" refer to conditions that vary by up to 20% (i.e., ±20%), especially up to 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% with respect to a particular condition (e.g., amount, level, concentration, time, etc.).

[0133] As used herein, the term "and / or" refers to all possible combinations of one or more of the associated listed items, and also includes the absence of a combination when interpreted alternatively (or).

[0134] The term "plurality" refers to a number greater than 1, up to 2 to 1×10 15 (or any integer therebetween), and more (2, 10, 100, 1000, 10000, 1×10 6 、1×10 7 、1×10 8 、1×10 9 、1×10 10 、1×10 11 、1×10 12 、1×10 13 、1×10 14 、1×10 15 etc. (and all integers therebetween)), etc.

[0135] The term "subject" is used to refer to an animal (including humans and non-human animals) to which the present invention can be applied. The term "user" is used to refer to a human who applies a device to a human or non-human animal. The subject and the user may be the same human subject, but this is not necessarily the case.

[0136] As used herein, "biological fluid" refers to any biological fluid of a subject, including, but not limited to, interstitial fluid (ISF), blood, saliva, tear secretions, milk secretions, nasal mucus secretions, tracheal secretions, bronchial secretions, alveolar secretions, gastric juice, gastric contents, glandular secretions, vaginal secretions, uterine secretions, prostatic secretions, semen, urine, sweat, cerebrospinal fluid, glomerular filtrate, liver secretions, bile, exudate, etc. Any of these are used in contact with the electrodes of the present invention.

[0137] Unless the contrary intention is apparent from the context of use, the terms "needle", "microneedle", and "wire" are used interchangeably. Each is functionally the same or similar and can be inserted into the subject's skin to contact the biological fluid.

[0138] The present invention is based, at least in part, on the inventor's discovery that an electrochemical aptamer-based (EAB) sensor device capable of detecting multiple analytes can be manufactured by assembling at least two working electrodes having different aptamer species. The different aptamer species can be configured to detect the same analyte or different analytes. Since the assembled working electrodes are in proximity to each other, they can be incorporated into a single device, and being in proximity, each working electrode can faithfully detect the amount of each analyte in the test sample. Since the assembled working electrodes are in proximity, it is possible to manufacture a small wearable device capable of monitoring the amount of clinically important analytes in real time. In such a wearable device, the electrodes can be wires, needles, or microneedles and contact the subject's ISF or other relevant bodily material. For in vitro detection of analytes, since the working electrodes are in proximity to each other, they can contact, for example, a single sample placed in a tube or a microplate well.

[0139] The assembly of the individual working electrodes is performed by the user (separate from the manufacturing facility) and can be immediately customized for a specific combination of target analytes. For example, hospital staff may assemble several working electrodes to provide a customized EAB sensor device for a specific patient. For example, a diabetic patient with a bacterial infection may need to be monitored for glucose, serum concentration of the antibiotic vancomycin, and markers of vancomycin-induced nephrotoxicity such as creatinine. Such an assembly can also be used for external testing (e.g., using a blood sample in a container) or in vivo real-time analyte detection.

[0140] However, in some cases, the assembly of the working electrodes can be performed at a manufacturing facility according to a custom order or according to a predetermined group of working electrode characteristics commonly used in a particular application. The assembly in that regard can be performed by human or robotic means.

[0141] In a working electrode assembly, the working electrodes can be maintained in a fixed mutual spacing relationship. A fixed mutual spacing relationship can also be provided with respect to non-working electrodes such as counter electrodes and reference electrodes. Such a relationship can be realized and maintained by attaching the electrodes to some kind of mount. In one embodiment, the mount is a one-piece structure and includes openings sized to snugly accommodate each working electrode. In other embodiments, each working electrode is embedded in a part of the mount, and a number of parts are joined together (e.g., by snap fits) to form an entire mount that holds a number of different working electrodes. In yet another embodiment, the electrodes are fixed to a rigid framework that can maintain the working electrodes in a fixed mutual spacing relationship. As yet another possibility, the mount is provided in liquid form, the working electrodes are disposed within the liquid, and then the liquid is converted to solid form (e.g., by polymerization or drying) to maintain the working electrodes in a fixed mutual spacing relationship.

[0142] The mount is typically non-conductive to prevent short circuits between the electrodes. When a conductive material is used, an insulator may be used to prevent short circuits. The mount may have a substantially planar surface. In some embodiments, the mount is rigid, semi-rigid, or at least partially rigid to facilitate penetration of the electrodes when applied to the skin. The mount may be partially flexible or semi-flexible so that, for wearable applications, the EAB sensor device can conform to the outer surface or contour of at least a part of the subject's body.

[0143] The mount can be made of or can include woven fabrics and non-woven fabrics including electronic fabrics, natural or synthetic fibers, natural or synthetic fabrics, silk, organic materials, natural or artificial composite materials including polymer materials, glass, ceramics including polymer ceramics, porous materials, polymers such as hard or semi-hard plastics, machinable polymers such as acrylic, polycarbonate, polyetheretherketone, or PEEK, synthetic polymers such as other plastics made from polymethylmethacrylate or acrylic glass, methacrylate, thermoplastic plastics and thermosetting plastics such as acrylic resin, polycarbonate, and polyetheretherketone, thermoplastic polymers such as polyethylene terephthalate, doped polymers such as polyacetylene, polypyrrole, polyindole, and polyaniline, intrinsically conductive polymers, metals such as aluminum, copper, gold (including colloidal gold), silver (including colloidal silver), chromium, platinum, titanium, metal alloys including stainless steel, carbon including colloidal carbon, carbon nanomaterials, carbon composite materials including carbon such as graphene and graphite, semiconductors such as silicon, germanium, gallium arsenide, doped semiconductors, and can be manufactured from or include organosilicates.

[0144] In the context of the present invention, the electrodes can be manufactured in various shapes and geometries, but the specific shape for transdermal applications is optimized to reliably penetrate the skin by breaking through the stratum corneum. The device can be configured to be pushed against the skin of the subject so that the electrodes can break through the stratum corneum and penetrate the skin layer. For non-human applications, the stratum corneum can be replaced with a similar layer or a dissimilar layer on the surface of the subject.

[0145] Generally, each electrode has a shape of a pointed structure protruding from the mount. Typically, the electrode extends substantially perpendicular to the mount.

[0146] The protruding structure of each electrode may be of any shape as long as it is needle-shaped. For example, the protruding structure may smoothly taper from the base to form a pointed tip (e.g., conical shape), or a plurality of sides may extend from the base and converge to form a pointed tip (e.g., pyramid shape or triangular prism), it may taper in only one dimension, it may have a base with a relatively constant diameter and a curved side, or it may be segmented to form a pointed tip (e.g., a part of a cylindrical shape). Typically, the pointed tip is sharp. The electrode may or may not change in shape along its length. Further, any edge or side of the shape may be chamfered, curved, or rounded.

[0147] In some embodiments, the shape is a cone, or a pyramid such as a triangular pyramid, square pyramid, or hexagonal pyramid. In other embodiments, the shape is a tetrahedron or a triangular prism. In further embodiments, the shape can take the form of a rocket, turret, arrowhead, spike, or spear.

[0148] It will be understood that a variety of other shapes can be used. For example, the shape is a circular or elliptical cylinder, which are truncated. Other shapes described herein may or may not be truncated. The term "truncated" as used in this context refers to a shape cut by a plane parallel to the base, which may be called a frustum shape, more specifically a frustum of a cone, or a shape cut at an angle to the axis of the electrode, which may be called an angled truncated shape. In the case of an angled truncated shape, the truncation angle with respect to the axis of the shape is at least about 50° and about 75° or less. In some embodiments, the truncation angle is about 55° to about 70°, about 55° to about 65°, about 50° to about 60°. In other embodiments, the truncation angle is about 50°, about 60°, or about 65°, or about 70°. In a particular embodiment, the shape of the electrode is a truncated cylinder with a truncation angle of about 60° with respect to its axis.

[0149] It will be understood that electrodes may be provided on the mount in the same shape or in different shapes. For example, the electrode can be formed as a plate or blade having a sharp edge.

[0150] The electrodes contemplated in the present invention can generally be classified into four types: solid, coated, dissolved, and hollow. However, it will be understood that the mount can be composed of combinations of these four types. For example, the mount can be composed of a combination of a solid electrode and a hollow electrode. For example, in order to function as a counter electrode or a reference electrode, it does not have to be a specific type.

[0151] For example, a hollow electrode generally includes a hollow interior defined by an inner wall having an opening at an end intended to contact a biological fluid. The hollow interior may or may not conform to the outer shape of the electrode. In some embodiments, the hollow electrode has a generally circular hollow interior, such as a bore hole. The diameter of the circular hollow interior can range from at least about 0.1 mm to about 5 mm. In some embodiments, the diameter of the hollow interior ranges from about 0.5 mm to about 1 mm. The opening to the hollow interior is preferably near the end, but may be on the upper surface of the hollow electrode.

[0152] The outer wall of the electrode can be configured to abut against the stratum corneum of the subject in order to control the depth of penetration into the skin layer of the subject. For this purpose, a shoulder or ledge may be provided on the outer wall.

[0153] The outer wall may have a smooth surface or a rough surface and can include surface features such as raised portions, etching, serrations, anchors, barbs, etc., which can assist in engaging with biological tissue after the electrode penetrates the stratum corneum and is fixed within the subject. The ability of the EAB sensor to remain in place is particularly beneficial to ensure continuous measurements over a long period at the same site within the subject. Further, restricting the location where the measurement is taken ensures more accurate longitudinal monitoring. In some embodiments, the EAB sensor is configured to remain in place for at least 1 minute, at least 1 hour, at least about 8 hours, at least about 18 hours, at least 1 day (about 24 hours), at least about 3 days, at least about 4 days, or at least 1 week. In some applications, it may be necessary or desirable to remain in place for up to 1 month.

[0154] The outer wall of the electrode may or may not have voids. In some embodiments, the outer wall of the hollow electrode is porous or has a porous layer, which increases the effective surface area of the electrode or allows the target of interest to enter the pores, while excluding one or more other targets or substances depending on the size of the target of interest. The diameter of the pores may be less than about 10 μm, preferably less than about 1 μm.

[0155] It will be understood that the size of the electrode and its placement on the mount may vary depending on the intended application.

[0156] The electrode can be at least as long as the thickness of the stratum corneum, penetrate the skin layer to a depth of at least 100 μm, and be positioned within the biological tissue to be in contact with the biological fluid of the subject. In some embodiments, the length is at least about 10% greater than the thickness of the stratum corneum, at least about 20% greater than the thickness of the stratum corneum, at least about 50% greater than the thickness of the stratum corneum, at least about 75% greater than the thickness of the stratum corneum, or at least about 100% greater than the thickness of the stratum corneum. In some embodiments, the length is less than about 1500 μm, less than about 1000 μm, less than about 750 μm, less than about 600 μm, less than about 500 μm, less than about 400 μm, less than about 300 μm, less than about 250 μm, greater than about 100 μm, greater than about 50 μm, greater than about 20 μm, or greater than about 10 μm. In other embodiments, the length is from about 100 μm to about 1000 μm, from about 200 μm to about 1000 μm, from about 500 μm to about 1000 μm, from about 750 μm to about 1000 μm, from about 800 μm to about 1000 μm, from about 900 μm to about 1000 μm, from about 100 μm to about 900 μm, from about 200 μm to about 900 μm, from about 500 μm to about 900 μm, from about 750 μm to about 900 μm, from about 800 μm to about 900 μm, from about 100 μm to about 800 μm, from about 200 μm to about 800 μm, from about 500 μm to about 800 μm, or from about 750 μm to about 800 μm. In other embodiments, the length is about 600 μm, about 750 μm, about 800 μm, about 900 μm, or about 1000 μm.

[0157] In some embodiments, the electrode has a hierarchical structure and thus not all have the same length. In such embodiments, the length of the electrode may be in the range of about 400 μm to about 800 μm.

[0158] The base width of the electrode may be less than at least about 50% of the length, less than about 25% of the length, less than about 20% of the length, less than about 15% of the length, less than about 10% of the length, or less than about 5% of the length. In some embodiments, the base width is at least about 100 μm but less than about 400 μm. In other embodiments, the diameter is about 200 μm or about 300 μm.

[0159] The diameter of the electrode may be less than at least about 50% of the length, less than about 25% of the length, less than about 20% of the length, less than about 15% of the length, less than about 10% of the length, or less than about 5% of the length. In some embodiments, the diameter is at least about 0.1 mm to about 5 mm or less. In some embodiments, the diameter is about 0.5 mm to about 1 mm.

[0160] In some cases, it may be desirable for one microneedle to penetrate deeper into the skin compared to another microneedle. Thus, the two microneedles may terminate at different distances from the skin surface or may terminate at different distances from the electrode mount. In some embodiments, the lengths of the two microneedles are different. In other embodiments, the lengths of the microneedles are the same and the mount is configured to axially displace one microneedle relative to the other microneedle. For example, the mount may be multi-level such that a first electrode extends from a first level and a second electrode extends from a second level.

[0161] The electrodes can be provided in various arrangements, and the number of electrodes provided on the mount depends on the available surface area. The mount can include up to about 100 electrodes. In some embodiments, the mount includes at least 2 to less than about 50 electrodes. In other embodiments, the mount includes at least 2 to less than about 30 electrodes. In still other embodiments, the mount includes at least 2 to less than about 20 electrodes. In still other embodiments, the mount includes at least 2 to less than about 10 electrodes.

[0162] Typically, the arrangement is of relatively low density. This is because it facilitates the penetration of the stratum corneum by the electrodes and can avoid potential problems of skin penetration due to high-density arrangements. In some embodiments, the mount is composed of at least about 4 electrodes / cm 2 , at least about 8 electrodes / cm 2 , or at least about 16 electrodes / cm 2 .

[0163] The electrodes can be arranged in pairs, in groups, or as a matrix. The paired arrangement includes an even number of electrodes. The group arrangement includes from 1 to about 5 groups, with each group including from about 4 to about 8 electrodes. The matrix arrangement can include an even or odd number of electrodes, and such an arrangement may or may not have the same number of rows and / or columns. In some embodiments, the electrodes are arranged in a matrix selected from the group consisting of 2×2, 2×3, 2×4, 2×5, 2×6, 3×2, 3×3, 3×4, 3×5, 3×6, 4×2, 4×3, 4×4, 4×5, 4×6, 5×2, 5×3, 5×4, 5×5, 5×6, 6×2, 6×3, 6×4, 6×5, and 6×6. In any arrangement, the electrodes can be arranged at intervals of about 5 mm, about 4 mm, about 3 mm, about 2 mm, less than about 1 mm, or about 0.5 mm and greater than about 0.1 mm from each other. The interval can be measured from the point from the center to the center of each electrode.

[0164] Similar to the mount, the electrodes can be made of any suitable material as described elsewhere in this specification.

[0165] Typically, the mount and the electrodes are manufactured from different materials.

[0166] The structure of the mount and / or the electrodes can be manufactured using any suitable technique. For example, in the case of a silicon-based electrode structure, this can be carried out using etching techniques. For example, an electrode structure of a polymer or plastic can be manufactured using additive manufacturing such as 3D printing, or molding including injection molding. Solid polymer electrodes are typically manufactured by molding methods such as injection molding and micro molding. Generally, in the injection molding process, the material is heated to the melting point Tm and then the processing temperature is adjusted to inject the material into the mold at a predetermined rate. Injection molding equipment can be expensive but is suitable for large-scale manufacturing. UV rapid prototyping is also a technique that can be used for manufacturing hollow polymer electrode structures. In this technique, a polymer structure is 3D printed by using a computer model to selectively polymerize a photocurable material by applying light. When electro-polymerized, the electrode structure is subjected to a washing step before curing. The electrode structure can also be manufactured using a microcomputer numerical control (CNC) micromachining method. In this manufacturing method, the mounting and shape of the electrodes, the spacing, the tip shape, and the selection of materials (such as soft metals like aluminum, 316L stainless steel, and copper, machinable polymers like polymethyl methacrylate, and ceramics, etc.) can be manufactured flexibly. The CNC-based manufacturing method is very cost-effective, highly accurately reproducible (for example, with an accuracy of 1 μm), has less residual material, and is suitable for automated manufacturing.

[0167] In the context of an EAB sensor, each working electrode is equipped with an aptamer that has specificity for a specific target analyte. Thus, when it is necessary to detect four analytes, at least four different working electrodes need to be assembled.

[0168] A working electrode can have at least one associated counter electrode and at least one associated reference electrode. Each working electrode can have a dedicated counter electrode, but in some embodiments, the counter electrode is shared by some or all of the assembled working electrodes. Each working electrode can have a dedicated reference electrode, but in some embodiments, the reference electrode is shared by some or all of the assembled working electrodes.

[0169] EAB sensors that are potentially useful in the context of the present invention are of the potential difference measurement type, current measurement type, or conductivity measurement type. In a potential difference measurement sensor, local equilibrium is established at the sensor interface, the electrode or membrane potential is measured, and information about the sample is obtained from the potential difference between two electrodes. A current measurement sensor applies a potential between a reference electrode and a working electrode to cause oxidation or reduction of redox-active species and measures the resulting current. A conductivity measurement sensor relies on the measurement of conductivity at a series of frequencies.

[0170] EAB sensors are typically of the current measurement type, and an aptamer (DNA, RNA, XNA, etc.) is bound to the working electrode. Gold is often used as the probe surface of the working electrode. The aptamer is accompanied by redox-active species that function as reporters. The redox reporter is often methylene blue. When a target (e.g., a drug) binds, the aptamer undergoes a structural change, bringing the redox reporter closer to the working electrode surface. This increased proximity results in an increase in electron transfer from the redox reporter to the electrode. The increase in the electron transfer rate contributes to the change in the Faradaic current detected by a potentiostat.

[0171] An aptamer is a small (usually 20 - 60 nucleotides) single-stranded RNA, DNA, or XNA oligonucleotide that can bind to a target drug with high affinity and specificity. An aptamer can be considered a nucleotide analog of an antibody, but the production of an aptamer is a much simpler and less expensive cell-free in vitro process compared to the production of antibodies by cell culture or in vivo methods.

[0172] An aptamer is usually selected from a combinatorial library containing a huge number (up to 10 18 ) of different oligonucleotides. RNA aptamers have significantly higher structural diversity compared to DNA aptamers, but their application is complicated due to problems with stability in the presence of RNases, high temperatures, and adverse pH.

[0173] The selection of aptamers that are selective for a given drug can be facilitated by a process called SELEX (Systematic Evolution of Ligands by Exponential Enrichment). This process can be considered in two alternating stages. In the first stage, a library of oligonucleotides is amplified to a desired concentration by polymerase chain reaction (PCR). In the selection of RNA aptamers, single-stranded oligoribonucleotides are generated by in vitro transcription of double-stranded DNA by T7 RNA polymerase. In the case of DNA aptamers, a pool of single-stranded oligodeoxyribonucleotides is generated by strand separation of the double-stranded PCR product. In the second stage, the amplified product is incubated with the target drug, and the oligonucleotides that bind to the drug are used in the next SELEX round.

[0174] The separation of oligonucleotides with high affinity for the target drug and the removal of unbound oligonucleotides are achieved by intense competition around the binding site. The selection pressure increases with each SELEX round. The maximum enrichment of the oligonucleotide pool containing the aptamer with the strongest affinity for the target molecule is typically achieved after 5 to 15 rounds.

[0175] EAB sensors are typically incorporated into a circuit that includes 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, and a point is formed at which the redox potential of the working electrode is measured relative to this. Also, the circuit typically includes a counter electrode that functions as the cathode or anode of the working electrode. Since the applied voltage bias does not pass through the reference electrode (due to the impedance of the potentiostat), the generated potential is due to the working electrode. The current is measured as the potential of the interrogating electrode relative to the stable potential of the reference electrode. A potential difference generates a current in the circuit, thereby generating an output signal. This signal ideally quantifies the target binding in response to electron transfer that is stoichiometrically proportional to the target binding.

[0176] The assembled device is particularly suitable for use as a wearable device, and can perform measurements while the subject is performing normal activities and / or over a long period of time. The wearable device may be a collar, bracelet or other suitable jewelry, watch, clothing, strap, adhesive, or patch. Those skilled in the art will understand that means for assisting in adhering and / or fixing the wearable device to the subject during use, such as micro-anchors, etc., may be provided.

[0177] The wearable device may include a housing structure that includes one or more other components such as an electronic processing unit. The electronic processing unit is configured to communicate electrically, either directly or indirectly, with at least one conductive element, and generally includes any one or more of a power supply, a data processing unit, an analog front end, and a wireless transmitter.

[0178] The housing structure is configured to at least partially cover the device, and the electrodes (such as micro-needles) protrude from the plane of the housing structure. The electrodes can be protected by a protective cover, and the protective cover can be removed before use to expose the protruding electrodes.

[0179] The device may further include means for monitoring the temperature or pH of the biological fluid if the effectiveness of the output depends thereon (such as the temperature or pH of the biological fluid), or if the operation or output can be adjusted.

[0180] The housing structure may be configured to cover and couple to the device by any suitable mechanism. For example, electromagnetic coupling, mechanical coupling, adhesive coupling, or magnetic coupling, etc. In some embodiments, the device and the housing structure can be detached by the coupling mechanism, making the housing structure and other components reusable, while the device can be discarded and replaced with another device as needed.

[0181] The wearable device may further include a computer program product executable as a software application resident on a mobile communication device that communicates with an electronic processing unit, the computer program product being capable of controlling one or more of (i) detection of an electrochemical measurement implemented on an electrode-based platform, (ii) data analysis, (iii) data transmission, (iv) device configuration, and (v) device power management. Examples of suitable mobile communication devices include, but are not limited to, smartphones, smartwatches, tablets, smart glasses, laptops, or other personal computers.

[0182] In some embodiments, the device itself includes a processor that includes program instructions configured to drive on-board functions such as voltammetry and transmit the output to a remote device via a wireless module such as a Bluetooth® module.

[0183] The present invention will be described in more detail with reference to the following non-limiting embodiments and drawings.

[0184] Referring to FIG. 1, exemplary means for the selection and assembly of a working electrode in the manufacture of an EAB sensor device are shown. In this example, the working electrode library has four electrode types 1, 2, 3, 4 (marked 10, 15, 20, 25 respectively). In practice, a much larger number of electrodes are provided in the library. Each electrode type has a plurality grouped together and held in a dedicated holder (30, 35, 40, 45 respectively).

[0185] A blank mount (50) is provided, on which a counter electrode (55) and a reference electrode (60) are already attached. The blank mount (50) further includes a series of openings (marked 65), each opening being sized to receive and hold one of the working electrodes (10, 15, 20, 25).

[0186] In the example of FIG. 1, it is desired to manufacture an EAB sensor device capable of detecting three different analytes. The first analyte is detected by the first electrode type (10), the second analyte is detected by the second electrode type (15), and the third analyte is detected by the third electrode type (25). The EAB sensor device need not detect other analytes.

[0187] In the assembly method, working electrodes of types 1, 2, and 3 (excluding type 4) are removed from their respective holders and inserted into the openings of the blank mount to produce the filling mount shown on the right.

[0188] Of course, in order to manufacture a functional sensing device, each working electrode in the filling mount is electrically connected to further hardware (not shown) configured to supply power and receive electrical signals from the electrode. The hardware can direct the supply of power (e.g., a potential that varies according to a square wave) and access software instructions that analyze the electrode output.

[0189] FIG. 2 shows the manufacturing method more extensively, and the steps of FIG. 1 are shown in the last two stages. The process of FIG. 2 begins with thousands of microneedles processed by various alternative means in various steps, as shown in FIG. 1, to provide various types of working electrodes.

[0190] Continuing with FIG. 2, all electrodes are subjected to surface treatment and cleaning. Either of two alternative surface coatings (A or B) can be applied (e.g., a specific metal coating). Thereafter, each of these A- or B-coated electrodes can be functionalized with an analyte-specific aptamer (type A, B, or C). Thereafter, a further passivation step (A or B) can be applied, followed by an essential sterilization step. The resulting electrodes (labeled "sensor type" in the figure) can be attached in various forms to the patch blank to provide a customized set of electrodes for analyte detection.

[0191] Figure 3 shows an exemplary solid micro-needle electrode (100) useful as a working electrode in the context of the present invention. When the working electrode is a micro-needle, the use of solid micro-needles provides advantages (compared to hollow micro-needles). Hollow micro-needles are difficult to manufacture on a large scale because they require drilling or injection molding of holes less than 0.3 mm, and it is difficult to achieve placement from the perspective of meeting a given tolerance range. Furthermore, the analyte detection surface area is relatively small because it is a subset of the inner diameter of the hollow micro-needle. Additionally, when the detection surface is recessed below the surface of the hollow needle, concentration changes need to be equilibrated by diffusion. Even a small gap can result in a long delay time. There is even a possibility that air can be trapped in this location, leading to equipment failure.

[0192] Continuing with Figure 3, the micro-needle electrode (100) has a tip (105) configured to pierce the skin of a subject.

[0193] Above the tip (105) is a detection region (110) coated with an analyte-specific aptamer that functionalizes the electrode to specifically detect a particular analyte.

[0194] Above the detection region (110) is an expansion region (115) that aids in handling, aligns the needle for correct insertion into a mount, and functions to abut against the mount after insertion.

[0195] Above the expansion region (115) is an electrical connection region (120) configured to be electrically connected to further hardware of the device, supply power to the electrode via an interface, and receive signals from the electrode. This interface preferably has low electrical and mechanical resistance, is easy to align, and can withstand a certain degree of bending and compression. Conductive and compressible foam, plastic, silicone, or epoxy can be placed independently on each needle or, in the case of anisotropy (Z-axis conductivity), can be placed once over all the assembled needles.

[0196] Dimensions (shown in mm) and angles are for illustration only and should not be regarded as limiting the electrodes depicted in FIG. 3.

[0197] Next, refer to FIGS. 4A and 4B showing embodiments of the functionalized microneedle (200) including the overmold portion (205). The overmold portion (205) is shown to extend through the lower part of the enclosure (300). The microneedle has an electrically insulating coating (210) that extends upward within the overmold (205).

[0198] Refer to the embodiments of FIGS. 4A and 4B in further detail. The overmold portion (205) has a support structure (marked as 215) that facilitates handling of the functionalized microneedle electrode (200) and insertion into the mount (400) through the opening (405). The overmold portion (205) and the mount (400) engage by press-fitting.

[0199] Upon press-fitting, the support structure (215) is removed by cutting at the level marked A - A'.

[0200] The press-fit itself functions as a waterproof seal and can prevent the intrusion of fluid (such as sample fluid or in-situ biological fluids like ISF) into the electrical connections and electronic devices above. Other or additional means for sealing will be apparent to those skilled in the art who benefit from this specification and include flexible seals, curable materials, and screw connections, etc.

[0201] In some embodiments, all of the assembled electrodes have a consistent base material and / or shape and / or dimensions. This is facilitated by large-scale batch production of the electrodes for subsequent functionalization with aptamers as needed. The metal composition and shape of the base electrode material can be adjusted to meet several requirements, such as the electrochemical surface area, surface roughness, retention within the mount by incorporation of inclined surfaces or facets (such as barbs), increase in surface area by macroscale shape modification (ridges, eyes, grooves, etc.), and separation of the surface area by incorporation of a dielectric masking layer.

[0202] Regarding the electrochemical surface hardness, the electrode can be composed of an alloy to increase the hardness and reduce the risk of damage to the electrode. The surface of the electrode can be plated with gold, and the underlying electrode is preferably an alloy containing cobalt, beryllium, or nickel.

[0203] When the working electrode is a microneedle configured to pierce the skin and contact the ISF or other biological fluids, its shape and / or material can be optimized for that purpose. Regarding the shape, the electrode can include chamfers (single or multiple), grinds (conical, undercut), or be hollow (some chemicals may benefit from the use of hollow needles). Regarding the material, the microneedle can be formed of a steel base layer plated / coated with gold or silver. In some embodiments, the needle is a solid material (including gold or silver). Additionally, the needle may be an alloy to increase rigidity or maintain sharpness.

[0204] When the EAB sensor device includes two or more working electrodes specific to the analyte and one reference electrode, and each working electrode is arranged at a substantially equal distance from the reference electrode, the detection of two or more target analytes can be improved. Additionally or alternatively, when the EAB sensor device includes two or more working electrodes specific to the analyte and one counter electrode, and each working electrode is arranged at a substantially equal distance from the counter electrode, the detection of two or more target analytes can be improved. In such an arrangement, when the EAB sensor device operates under voltammetric conditions, undesirable peak splitting and / or peak broadening are avoided. As a result, a single peak output and / or a narrow peak on the voltammetric curve are obtained, and such an output improves the detection of the binding of the target on the working electrode coated with the aptamer.

[0205] In some applications, non-equidistant arrangements may be preferred, in which case a deviation of at least about 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100% from the equal distance can be implemented. Such applications may occur when peak splitting or broadening is not a substantial problem and flexibility regarding the spatial arrangement of the electrodes is required. For example, the shape, dimensions, or arrangement of the device may necessitate a specific method that does not allow the use of equidistant electrodes.

[0206] The assembled device of the present invention can be operable by voltammetry including square wave voltammetry, cyclic voltammetry, chronoamperometry, chronopotentiometry, impedance spectroscopy, electrochemically implemented surface plasmon resonance, waveguide optical mode spectroscopy, ellipsometry, and quartz crystal microbalance, or field effect transistor-based methods. Generally, the EAB sensor device is configured to be operable by voltammetry (especially square wave voltammetry), and in that regard, it is configured to be connected to a voltage source, especially a controllable voltage source. When at least both the working electrode and the counter electrode are in contact with the biological fluid, a voltage is applied to at least the working electrode and the counter electrode.

[0207] The microneedles may have other features to facilitate assembly. Such features include a scaffold that can be removed before or after assembly to facilitate handling, an increase in size to simplify the handling or placement of the needles, incorporating a flat surface, groove, or keyway into the needle design to improve assembly accuracy, using overmolded or insert-molded plastic around the electrode needles, and the like.

[0208] In addition to the advantages described above, the present invention provides advantages in the process for manufacturing sensor devices. Working electrodes specific to a number of individual analytes can be prepared in batch numbers of at least about 10,000, 50,000, or 100,000. There is a technical advantage in that the larger the batch, the less often the manufacturing conditions need to be optimized.

[0209] Economic advantages are also obtained from the scale and the low frequency of execution of the QA / QC process. The individual electrodes of the batch can be incorporated into sensor devices as needed. In this way, the manufacturer can provide the desired combination of working electrodes in response to consumer demand, thereby preventing the accumulation of unwanted combinations in inventory.

[0210] Independent assembly of electronic and mechanical components. Inserting the assembled independent needles into the bottom of the device enables asynchronous construction of the device and the electrodes. Thereby, the device can be assembled, sterilized, and configured independently of the sensor. This is preferable because it limits the exposure of the sensor to oxygen and may require special sterilization techniques. Furthermore, the working electrodes can be selected and a large number of devices can be manufactured and stockpiled by a contract manufacturer at another facility or before final configuration.

[0211] As a further advantage, the same mechanical and electrical components can be used interchangeably between a plurality of different sensor types, reducing development costs and time. Further, since the sensor device can be customized for the analyte to be detected later, a large single batch of mechanical and electrical components can be prepared.

[0212] The functionalized surface of the working electrode is sensitive to physical wear. Using mounts or dielectric masking, the components can be mechanically connected without contacting the functionalized surface.

[0213] Regarding tolerances during high-speed and mass production, further advantages can be found. In the manufacture of electrodes, it is difficult to maintain very accurate and precise tolerances (<+ / -25μm) for mass production and high speed. By using larger electrical connections and press-fits to allow for some variation in component position, the tolerance requirements are reduced to a value closer to + / -100μm compared to other configurations (i.e., hollow microneedles).

[0214] The method includes assembling two or more electrodes with a device for contacting the electrodes with the skin of a subject. The electrodes (which may be microneedles) may be attached to a movable part of the device or to another part of the device. In some embodiments, the movable part moves in a non-linear path.

[0215] The non-linear path may be of limited length and the path may be an arc of limited angle. With this arrangement, the main movable part of the device only requires a limited movable range in the vertical direction to press and insert the microneedle into the skin of the subject. Due to the limited movable range, when viewed from the side, the housing of the device can have a relatively low profile. Thus, the device rises to a relatively low height above the skin and is therefore less obtrusive to the subject.

[0216] Furthermore, due to the non-linear path of the movable part, a simplified mechanism can be used. For example, the movable part can be moved by a simple bending mechanism or a hinge mechanism. Since these mechanisms require relatively few parts, it is possible to develop a device that is smaller, lighter, simpler, more reliable, and less expensive overall.

[0217] Certain embodiments of the present invention have additional features that, alone or in combination with other features, provide further advantages over the prior art or further useful alternatives. Such embodiments will be described more fully with reference to the non-limiting preferred embodiments described below.

[0218] Referring to FIG. 5, a basic form of the device (500) is shown, including a microneedle array (marked with one microneedle (515)) attached to the movable part. In this embodiment, the movable part is an elastically deformable arm (520). The arm (520) is biased to take a linear configuration (520b), but is first presented to the user as an upwardly bent arm (520a) as shown by the dashed line.

[0219] The device (500) includes a rigid housing (525) having a skin contact portion (530) on the lower side and defining a downward skin contact surface (535). The surface (535) is placed on the skin of the subject and held to the subject's skin by areas of dermatologically acceptable adhesives (540a, 540b). Suitable adhesives can typically be made water-resistant so that the subject can bathe as normal. The adhesives typically have sufficient adhesive strength to prevent peeling that may occur during daily activities such as changing clothes, undressing, sleeping, doing housework, light to moderate intensity sports activities, and brushing against objects while walking. The level of the adhesive is typically not so high as to cause difficulty, discomfort, pain, irritation, or skin damage when removing the device.

[0220] Exemplary adhesives are synthetic rubber adhesives or pressure-sensitive acrylic adhesives of the type used in medical tapes. A double-sided medical tape such as 3M (registered trademark) 1577 tape may be used to adhere one side to the device and the other side to the skin of the subject.

[0221] The skin contact portion (530) includes a space (545) marked with marks (545a) and (545b) at the edges. The space (545) provides a passage for the microneedles (515) to pass through and allows the distal region of the microneedles to penetrate and embed into the underlying skin (550) when the arm (520) is in the straight position (520b).

[0222] The arm (520) is held in a bent state by a shelf (555) that functions as a release means. When the user wants to insert the microneedles (515) into the skin (550), the button (560) is pressed as indicated by the arrow. The lower surface of the button (560) abuts against the shelf (550), and since the shelf (555) has the ability to deform (e.g., is made of a material such as rubber or is formed from flexible protrusions on the inner surface of the housing (525)), it bends downward under the applied force and releases the edge of the arm (520a). Due to the elasticity of the arm (520a), the arm rapidly returns to the biased straight position (520b), thereby pushing the microneedles (515) into the underlying skin (550). The shelf (555) is configured to exhibit sufficient elasticity to resist the biasing force of the arm (520a), but its elasticity is not sufficient to resist the downward force exerted when the button (555) is pressed.

[0223] In the embodiment of FIG. 5, one end of the arm (520) is fixed to the housing (525) by a fixture (565). The arm (520) is flexible, but not flexible enough to easily move away from the position (520b) when placed on the skin (550) of the subject. As can be understood, when the arm (520) moves away from the position (520b), the microneedle (515) can be withdrawn from the skin (550). Considering that the arm (520) is biased towards the position (520b), a locking mechanism may not be necessary to maintain the arm in the position (520b). However, if necessary, for the embodiment of FIG. 6A, an appropriate locking mechanism will be described below.

[0224] FIG. 6A shows an alternative basic form of the device (500), where the arm (505) is rigid and is hinge-coupled to the housing (525) by a hinge pin (510). The embodiment of FIG. 6A operates in the same manner as the embodiment of FIG. 5 as long as the shelf (555) acts as a release means. However, in the embodiment of FIG. 6A, the button (515) acts on the rigid arm (505a). The rigid arm (505a) transmits the force of the button to the deformable shelf (555), and by bending the shelf (555), the free end of the arm (505a) is released. The button (515) is continuously depressed by the user until the arm takes the position (505b), at which position the microneedle (515) is embedded in the skin (550). Also in this case, the point at the free end of the arm (505a) moves along a non-linear path, and in this embodiment, the path is an arc that is part of a circle, and the origin of the circle is at the hinge pin (510).

[0225] In the hinge arrangement of the embodiment of FIG. 6A, it can be seen that while the device is being worn, there is no resistance to the hinge movement of the arm (505) away from the position (505b). Therefore, there is a risk that the micro-needle (515) will remain withdrawn from the skin (550). Accordingly, a locking mechanism is provided to maintain the arm in position (505b). This mechanism includes, for example, a deformable latch (520) made from a material with some flexibility or from internal protrusions formed from the housing (525) material. The latch (520) has an inclined upper surface, and when it contacts the rigid arm (505), the entire latch (520) bends to the left (as shown in the figure) by the force applied by the user through the button (515) and the inclined upper surface. When the end of the arm (505) passes through the lower corner of the inclined upper surface, the latch (520) returns to its normal upright position (as shown in the figure), and the free end (505b) of the arm is securely fixed in the recess at the base of the latch (520).

[0226] An alternative to the embodiment of FIG. 6A is shown in FIG. 6B. In FIG. 6B, the device (500) does not have an upper housing. The arm (505a) is maintained in a predetermined position by release means (555), but in this embodiment, this release means can be removed by the user when the device (500) is applied to the subject. After removing the release means (555), the arm (505a) is pushed downward by the user to take a second position (505b).

[0227] In the embodiments of FIGS. 5, 6A, and 6B, it is noted that the free end of the arm (520 or 505) moves non-linearly when returning to the biased position when released from the shelf (555). Considering a point on the free end of the arm (520 or 505), that point moves along a non-linear path that describes an arc. In the context of the present invention, the terms "arc", "bow", and similar terms refer to a curve connecting any two points. The term "arc" should not be construed restrictively to mean only a segment of a circle, but in some embodiments, it is a segment of a circle (see, for example, the embodiment of FIG. 6A).

[0228] As is apparent from both the basic embodiments of FIGS. 5 and 6A and 6B, in either case, the arm (520 or 505) moves a relatively short distance when transitioning from the first position to the second position. In fact, in these embodiments (and other specific embodiments), the device is intentionally configured such that the arm cannot move along a path outside the path between the first and second positions. In other words, the device may be configured such that the arm cannot move along a path outside the shortest distance between the first and second positions.

[0229] By restricting the path along which the arm can move, the advantage is obtained that the height of the device (the vertical direction shown in the figures) is also restricted. Thus, the device can have a low profile that extends upward from the skin of the subject by a relatively short distance (in terms of dimensions).

[0230] Next, referring to FIGS. 7A and 7B, a preferred device is shown that is configured substantially in accordance with the embodiment of FIG. 5 and is operable in concert with the embodiment of FIG. 5. The arm (520) is integrally formed with a PCB (565) carrying various electronic components necessary for the operation of the device. The PCB material is elastically deformable, and the arm (with micro-needles attached to the terminals) bends upward when pulled out, positioning the arm in the first position, but when released, it assumes the second position due to the natural biasing of the arm towards the second position.

[0231] The arm (520) is maintained in the first position because the end of the arm (520) rests on the shelf (555), as most clearly shown in FIG. 5A. In this position, the micro-needles (not shown, extending downward from 570) are held within the device without penetrating the space (545). This is the configuration in which the device is provided for use and applied to the skin of the subject.

[0232] The arm (520) is connected to a micro-needle mounting block (570) that supports the micro-needles. The mounting block (570) also includes conduits (not shown) for carrying current from each of the micro-needles (515) to one of a number of connection points (575) on the PCB (565). This arrangement allows electrical signals to be transmitted to or from the micro-needles embedded in the subject's skin. For example, the device can be configured as a sensor that includes micro-needles configured to contact a biological fluid in the subject's body and detect an analyte therein. The biological fluid can be, but is not limited to, interstitial fluid, blood, or a mixture thereof. The electrical signals from the micro-needles are transmitted to the PCB for amplification, filtering, encoding, analysis, transmission, or other electrical or electronic processes.

[0233] In this embodiment, the PCB serves a dual function of housing the electronics of the device and as a driving means for moving the micro-needles from an internal position of the device to an external position. The PCB material has been found to be very suitable for providing a limited but preferred range of movement for the arm of the device. This arrangement reduces the number of components within the device.

[0234] On the upper surface of the housing (525), the operating surface of a button (516) that can be pressed by the user's finger is exposed. The button (516) is biased upward (as shown in the figure) by a spring or is integrally formed with the housing (525) material. In the latter biasing form, the button (516) is attached to an arm integral with the housing material and is biased such that the upper surface of the button (516) is coplanar with the housing (525).

[0235] The lower part (not shown) of the button (516) abuts against the upper surface of the arm (520), and the upper surface is the back surface of the PCB (565). When the button (516) is pressed, the arm (520) is pushed downward and disengages from the shelf (555), taking a second position. In the second position, it can be seen that the microneedles extend through their respective spaces (545) and are implanted into the underlying skin (e.g., the epidermis, dermis, or subcutaneous tissue of the subject).

[0236] The natural biasing of the PCB (565) material towards the second position is strong enough to hold the arm in the second position without the need for means to lock the arm (520) in the second position. Thus, the microneedles can remain implanted in the subject's skin for an extended period.

[0237] In an alternative embodiment, the arm (520) has a curved configuration when in the second position and is naturally biased in a direction away from the second position. In another embodiment, the biasing of the arm (520) towards the second position is not strong enough to prevent movement away from the second position. In such embodiments (and other embodiments), a locking mechanism is provided to prevent the arm from moving away from the second position so that the microneedles do not retract into the device and remain implanted in the skin. A suitable locking mechanism is the latch mechanism disclosed in connection with other embodiments of this specification. Other locking mechanisms will be apparent to those skilled in the art who benefit from this specification.

[0238] The housing (525) includes opposing recesses (517) for gripping between the user's thumb and index finger and facilitating pressing the device against the skin surface. The user's index finger can freely actuate the button (516) to implant the microneedles under the skin.

[0239] A dermatologically acceptable adhesive layer (not shown) may be applied to the skin contact surface (535) so that the device can be maintained in place on the subject's skin over a long period of time. The adhesive layer can cover part or substantially all of the skin contact surface (535). As described in other embodiments of the devices herein, a manually removable flexible layer can cover the adhesive until the device is applied to the skin.

[0240] Next, referring to FIGS. 8, 9A, 9B, 10, and 11, a preferred device is shown that is configured substantially in accordance with the embodiment of FIG. 6B and is operable in a substantially consistent manner.

[0241] This embodiment includes an upper housing portion (525) and a skin contact portion (530). A removable flexible layer (590) that can be grasped via a tab (595) is also provided, and removing this exposes a dermatologically acceptable adhesive on the skin contact surface. As described above, the adhesive is for holding the device on the subject's skin over a long period of time. The flexible layer (590) serves to prevent curing or drying of the adhesive, contamination of the adhesive layer prior to use, and / or premature adhesion of the adhesive to packaging or other surfaces. In a particularly preferred embodiment, the flexible layer (590) in addition to covering the adhesive layer extends over the space (545) to prevent contamination of the microneedles and also helps to prevent unintentional needle stick injury to the user.

[0242] The device may have a holding portion that functions to hold the device on the skin such that the protruding portion remains in contact with the subject's body fluid. The holding portion may be dedicated to that function or may serve another function.

[0243] In many cases, an adhesively coated backing or a skin-scientifically acceptable adhesive may be useful. The adhesive simplifies the application of the device by the user, and in many cases, it is only necessary to peel off the protective backing to expose the adhesive and then bring the exposed adhesive into contact with the skin. Since this application method is similar to that of a bandage, it is a process already familiar to the user.

[0244] Instead of using an adhesive, the retaining portion may be any mechanical means for maintaining the device in the required position on the skin. For example, the device may include a dedicated strap that engages around the limb and can be adjusted to keep the device firmly applied to the subject. As an alternative, the device may be incorporated into a wearable item such as a glove or a shirt, or a jewelry item such as a ring that serves to hold the device in place. The device may be configured to engage with an individual wearable item (e.g., by complementary hook and loop means), and the wearable item may be integrated with the device.

[0245] In some embodiments, the device is held only by a wearable item that abuts against the housing. For example, the retaining portion may be a stretchy glove that fits snugly over the device.

[0246] In some embodiments, the retaining portion is the surface or part of the device that contacts the subject's skin, and the characteristics of the subject at least partially play a role in maintaining the device in a predetermined position on the subject. For example, the device may be configured to be held between two parts of the body that are normally closely juxtaposed, or to be held within an existing anatomical structure. The device may be shaped and / or dimensioned to be held between the toes, on the buttocks, in the groin, in the mouth, in the nostrils, in the ear canal, or at the navel.

[0247] In other embodiments, the device housing is shaped and / or sized to fit snugly, for example, on a finger, a toe, or an ear. The device housing is elastically deformable and may be made of, for example, a rubber material and configured to be stretched over any anatomical part (such as a finger).

[0248] Each of the foregoing embodiments is considered to be a holding part in the context of the present invention.

[0249] The device further includes a release member (600) having a gripping portion (605) and a wedge portion (610), the function of which will be described in more detail below.

[0250] Next, referring to the exploded views of FIGS. 9A and 9B, components similar to those in the previous figures will become immediately apparent.

[0251] In this embodiment, the motive force for moving the arm (505) and thereby pushing the microneedle (515) into the underlying skin is provided by the user. In use, the user places a finger on the upper housing (525) and presses downward. Further, the arm (505) is movable by a hinge configuration.

[0252] The hinge configuration is provided by opposing protrusions (715) extending from the skin contact portion (530), each protrusion including an opening. The arm (505) includes opposing laterally extending disks (571), each disk fitting into the opening of a protrusion (572). It is clear that the arm (505) can be hinged open and closed relative to the skin contact portion (530), allowing movement from a first position to a second position.

[0253] The arm (505) is presented to the user with the arm in the first position. The arm (505) is maintained in the first position by the wedge portion (610) of the release member (600). Before removing the release member (600), the wedge portion (610) is inserted between the skin contact portion (530) and the arm (505), thereby holding the microneedle within the device.

[0254] When attempting to apply the device to the skin of a subject, the user pulls on tab (595) to remove the flexible layer (590), exposing the adhesive layer of the skin contact surface (535). Next, the device is applied to the skin, and the adhesive keeps the device held in place for an extended period of time.

[0255] Once the device is applied to the skin, the user grasps the gripping portion (605) and pulls it laterally to the left (as shown in the figure), completely removing the release member (600). Since the release member (600) no longer has any function, it is discarded at this point. Removing the release member (600) releases the arm (605) from its first position and allows it to move to a second position (by the downward force applied by the user), where the lower surface of the arm (605) contacts the upper surface of the skin contact portion (530). In the second position, the microneedles (515) extend through space (545) and into the skin below.

[0256] As can be appreciated, the release member (600) can be configured to prevent the upper housing (525) of the device from closing (approaching) the skin contact portion (530) when not intended by the user. The release member (600) is inserted between the upper housing (525) and the skin contact portion (530) or otherwise juxtaposed such that the upper housing (525) is prevented from closing towards the skin contact portion (530) enough for the tip of the microneedle (i.e., the protruding portion) to protrude from the base of the hole in the skin contact portion (530). Preventing closure also prevents the arm (505) from moving from its first position to its second position. Thus, when the release member (600) is in place, inadvertent access to the tip of the microneedle does not cause contamination or damage to the microneedle. When using the device, the user removes the release member (600) as a step in the usage process. In a preferred embodiment of using the device, the user first attaches the device to the skin of the subject, then removes the release member (600), and then presses on the upper housing (525) to insert the microneedles into the skin.

[0257] Before being removed by the user, the release member (600) can be held in a predetermined position by any of various features. In one example, the release member (600) includes protrusions that fit into recesses in the upper housing (525), the skin contact portion (530), or both the upper housing (525) and the skin contact portion (530) to assist in holding the release member in a predetermined position until it is intentionally removed. In another example, the release member (600) is designed to be slidably assembled to the skin contact portion (530) or the upper housing (525), and the friction between the release member (600) and the upper housing (525) or the skin contact portion (530) helps to hold the release member in a predetermined position until it is intentionally removed. In yet another example, a magnetic force can be used to assist in holding the release member (600) in a predetermined position. In one embodiment of the present invention, a magnet mounted within the release member (600) is positioned such that when the release member (600) is in a predetermined position, it is proximate to a Hall effect sensor positioned in either the upper housing (525) or the skin contact portion (530). According to this embodiment, when the release member (600) is removed by the user, the Hall effect sensor detects the removal of the magnet and causes the device to perform some action, such as powering on the electronic circuit to make it usable or converting from a sleep mode to an active mode. The above are examples of possible ways to assist in holding the release member (600) in a predetermined position before intentional removal, which can be used alone or in combination, and it should be understood that other methods known in the art can also be used alone or in combination with the above examples.

[0258] In some embodiments of the present invention, the release member (600) can also function as a cover element used to cover the microneedle after the device is removed from the subject. In a preferred example of this embodiment, the locking element is on the upper housing (525) and extends downwardly towards the skin contact portion (530). The release member (600) includes a groove that allows the release member (600) to slide past the locking element when the release member (600) is withdrawn from the device, while the surface of the release member (600) facing the upper surface of the skin contact portion (530) is continuous. In use, the release member (600) according to this preferred embodiment is removed and held by the user before pressing the upper housing (525) to insert the microneedle into the subject's skin. After removing the device from the subject after use, the user is instructed to adhere the release member (600) to the adhesive layer on the lower surface of the skin contact portion (630) to cover the protruding microneedle. In another example of this embodiment, the release member (600) is flexibly attached to the device, and the release member (600) remains attached to the device even after the user has withdrawn the device, and can be repositioned to cover the protruding microneedle after removing the device from the subject after use. In yet another example of this embodiment, the release member (600) and the upper housing (525) are designed to be slidably or otherwise engageable with the upper housing (525) after the release member (600) is removed, and the release member (600) is intended to be stored during use of the device and removed for use as a cover element after removing the device from the subject.

[0259] In some embodiments, the device is configured to facilitate removal of the device from the subject. As will be appreciated, the use of an adhesive layer may make it difficult to remove the device from the skin. Examples of such configurations include leaving a portion of the skin contact surface (535) uncoated with adhesive, creating a gap between the subject's skin and the surface (535). In this case, the user can use this gap as a leverage point to assist in breaking the adhesive bond and pulling the device away from the skin. In another example, a lever mechanism is incorporated that is not located on the skin contact surface, allowing for a higher gap than the gap created by the absence of adhesive on a portion of the skin contact surface. In yet another example, a tab extending beyond at least one edge of the skin contact portion (30) and attached to the adhesive layer can be incorporated, in which case the user can pull on the tab with sufficient force to stretch and flex the adhesive layer and further peel the adhesive from the skin contact surface (535) and the skin.

[0260] In some embodiments of the present invention, the device is designed such that the release member (600) is locked in position prior to use of the device unless pressure is applied to the upper housing (525). This embodiment is intended to further reduce the risk of the release member (600) being prematurely withdrawn. In an example of this embodiment, the release member (600) and at least one of the upper housing (525) and the skin contact portion (530) have features that engage lockably when the upper housing (525) is not being pushed. When the upper housing (525) is pushed, at least one feature of the upper housing (525) and the skin contact portion (530) is deformed, releasing the engagement of the release member (600) and allowing it to be withdrawn.

[0261] In yet other embodiments, the release member (600) need not be removed by the user from the device. According to these embodiments, the release member (600) includes a flexible element having a rigidity high enough that it will not substantially flex even when subjected to the closing forces that may be applied by the user during manufacturing, storage, and prior to application to the subject, but is flexible enough that it will flex when the user intentionally applies a closing force to the device when applied to the skin of the subject. When flexed in this manner, the release member (600) flexes and the upper housing (525) can close towards the skin contact portion (530). In these embodiments, the release member (600) can also function as a locking element, or the release member (600) can be separate from the locking portion. In some of these embodiments, a structure such as that labeled (650) in FIGS. 9A, 9B, and 11 forms the release member (600).

[0262] Each space (545) of the device is sized such that the microneedles can clearly pass therethrough and is dimensioned such that at least the tapered portion of the microneedles does not strike the side of the hole during insertion. In some embodiments, the hole may have a sufficient cross-sectional area such that no portion of the microneedles contacts the side of the space during insertion. In other embodiments, at least a portion of the cross-sectional area along the length of the hole has a cross-sectional area such that a portion of the length of the microneedles contacts the side of the hole during insertion. According to this embodiment, the hole functions to assist in supporting a portion of the length of the microneedles such that it helps to prevent the microneedles from bending during insertion.

[0263] In some embodiments of the device, the skin contact portion (530) includes an additional space or recess configured to receive a protrusion of the release member and assists in retaining the release member until it is removed by the user. Additionally or alternatively, the skin contact portion (530) includes a protrusion designed to be received in a recess of the release member and assists in holding the release member in a predetermined position until the user intentionally removes it.

[0264] The embodiments shown in FIGS. 8, 9A, 9B, 10, and 11 include a locking portion in the form of a latch (650), which permanently locks the arm (505) in a second position and prevents the arm (505) from hinging. In the illustrated embodiment, the latch (650) can flex as the arm (505) moves toward the closed position, but returns to its original position when the arm (505) is in the second position (505b), thereby locking the arm (505) in a predetermined position, and is a simple one-piece member.

[0265] The locking portion may act not on the arm (505), but on another part of the device to lock that part in a position that locks the arm. For example, the locking portion may act on the upper housing (525), and the upper housing (525) may hold the arm (505) in the second position. As yet another alternative, the locking portion may act on the PCB (565), and the PCB (565) may hold the arm (505) in the second position.

[0266] In other embodiments, the locking portion includes a recess into which a protrusion of the upper housing (525) is inserted, locking the upper housing (525) in the closed position (i.e., when the arm (505) is in the second position). In one embodiment, the locking portion includes a flexible element designed such that the locking portion can move when the upper housing (525) hits it, so that the housing (525) can close against the skin contact portion (530), and when the upper housing (525) closes, the locking portion can move to lock the upper housing (525) in the closed position. In one embodiment, the device includes a protrusion on the upper housing (525) designed to be inserted into the recess of the locking portion, the protrusion including a flexible element to allow the protrusion to move, whereby the upper housing (525) can close against the skin contact portion (530), and then when the housing (525) closes against the skin contact portion (530), the protrusion moves and is inserted into the recess of the locking portion, locking the upper housing (525) in the closed position. The flexible element can include a shaft that is sufficiently deformable so that the upper housing (525) can be closed without the shaft yielding, such that the flexible element attempts to return to its original position after the upper housing (525) is closed. Although less preferred, in still functional embodiments, the flexible element includes a coil spring.

[0267] The flexible element of the locking portion can be manufactured from any suitable material having the required rigidity and yield point. Examples of suitable materials include amorphous plastics, crystalline plastics, spring steel, non-spring steel, stainless steel, or other materials known in the art having suitable mechanical properties.

[0268] In a preferred embodiment of the present invention, the locking portion is manufactured from the same material as the skin contact portion (530), facilitating the manufacture of the skin contact portion including an integral locking portion.

[0269] In a particularly preferred embodiment of the present invention, the force required to bend or otherwise move the flexible element is designed to be large enough such that the pressure that the user needs to supply to deform the flexible element and close the upper housing (525) towards the skin contact portion is sufficient to insert the microneedle into the skin. According to this embodiment, the flexible element of the locking portion is used to set the force required to close the device (thereby causing the arm to assume a second position), and that force is ensured to be sufficient to insert the microneedle into the intended position embedded in the skin.

[0270] In other embodiments, the locking portion includes at least one adhesive region located on at least one of the lower surface of the upper housing (525) and the upper surface of the skin contact surface (535). When the device is closed, the one or more adhesive regions adhere the upper housing (525) to the skin contact portion (530) and lock the device in the closed position.

[0271] In another embodiment of the present invention, the locking portion can take three different stable states. In the first state, the locking portion is in a disengaged configuration before the upper housing (525) is pushed downward toward the skin contact portion (530) to close the device. In the second state, the locking portion is in the first engagement position. When the locking portion is in the first engagement position, the locking portion functions to lock the microneedle (515) in the implanted position in the skin (i.e., the arm (505) is in the second position). In the third state, the locking portion is in the second engagement position. In this state, the locking portion locks the device in the open position (i.e., the arm (505) is in the first position), the microneedle is retracted into the device, and the risk of needle stick injury due to the microneedle protruding after device use is reduced. In an example of this embodiment, the locking portion includes a user engagement portion that can be grasped or otherwise engaged by the user. For example, by engaging the fingernail under the protruding shelf, the user can flex the flexible portion of the locking portion. According to this example, to close the device, the user presses the upper housing (525) to lock it in a predetermined position, similar to other embodiments disclosed herein. When it is necessary to remove the device from the subject, the user engages the locking portion and flexes it in a first direction to unlock the upper housing (525) from the skin contact portion (525), and then flexes the locking portion in a second direction to lock the device in the open position (i.e., the arm is in the first position) with the microneedle in the retracted position. In a preferred embodiment of this example, in the first direction, the locking portion moves away from the body of the device, and in the second direction, it moves toward the body of the device. When sufficiently biased in the second direction, the locking portion is designed to stably engage, for example, in a recess, preventing the device from closing even if not deliberately closed.

[0272] In some embodiments of the present invention, the downward force on the microneedles inserted into the skin is provided via a flexible element of a locking portion that applies a downward force when the device is locked in the closed position (i.e., the movable arm is in the second position). In some embodiments, effective locking of the movable arm in the second position is provided by a dedicated spring or other suitable biasing means. In other embodiments, the spring or other biasing means is not dedicated to the locking function and can also function, for example, as a driving force when moving the arm from the first position to the second position. For example, a torsion spring can apply a closing torque at the pivot point (if present). In yet another example, a flat, disk-shaped, or coiled spring is attached to the back of the microneedle, and the spring is deformed or compressed when the device is closed, and a downward force is applied to the microneedle when the device is in the closed position.

[0273] Although not an essential feature of the present invention, in many applications where the microneedles are used for the purpose of conducting an electric current to, from, or through the skin, a PCB (565) is required. In that regard, the PCB can carry a microprocessor and / or volatile electronic memory (such as RAM) and / or non-volatile electronic memory (such as ROM) and / or a wireless network module (such as a Bluetooth® module). Of course, the device typically includes a power source by a button cell.

[0274] The embodiment shown in FIG. 7A further includes a light-emitting diode (LED) (800) visible to the user. One function of the LED (800) can be to confirm for the user and / or the subject that the microneedles are properly embedded in the skin upon application and that their state of being worn over a long period is maintained.

[0275] The LED is electrically connected to a printed circuit board (PCB) (565), and the PCB is electrically connected to micro needles (515). Whether the micro needles are properly embedded can be determined by referring to any one or more of the current flow, resistance to current, or impedance between two micro needles.

[0276] Alternatively, proper embedding of a single micro needle can be determined by referring to any one or more of the current flow, resistance to current, or impedance between the single micro needle and other electrical contacts of the device and the skin. As an example, a conductive pad can be placed on the surface of the skin, and in some examples, the conductive pad is placed on the surface of the housing that contacts the skin. This conductive pad, in cooperation with at least one micro needle, completes an electrical circuit when the micro needle is inserted into the skin. Completion of this circuit is used to indicate proper insertion of the micro needle.

[0277] The associated electronic device is simple. As an example, a biological fluid of the skin, such as interstitial fluid (naturally conductive), functions to complete a circuit including the LED. Proper embedding is assumed to be indicated by the micro needle simply contacting the biological fluid. The LED lights up at the location where the micro needle contacts the biological fluid (or vice versa), thereby visually indicating proper embedding.

[0278] More advanced electronic arrangements may be required to more reliably ensure that the micro-needles are correctly embedded. For example, by considering whether the minimum length of the micro-needles is embedded, it is ensured that the micro-needles are inserted to a specific minimum depth. The device includes electronic means for measuring a quantity of a parameter such as the flow of current, and the greater the flow of current, the more completely the micro-needles are embedded. Program instructions executed by a processor incorporated in the device or associated with the device may use a parameter such as the flow of current as an input (and may also be used in combination with other physiological or environmental parameters) to provide an indication of the embedding of the micro-needles.

[0279] A further function of the LED is to provide other information such as the charge level of the battery. For example, the LED is connected to a microprocessor that can monitor the battery voltage, and when the voltage drops below a predetermined threshold, the microprocessor causes the LED to blink red. The value may be a voltage somewhat higher than the minimum operating voltage in order to give the subject time to access a replacement battery (or a replacement device if the battery is not repairable by the user) before the device becomes inoperable.

[0280] In other embodiments, the LED may generate an output indicator of the data connection state. For example, the LED may alternately flash red and green lights to warn of an interruption in the wireless data connection with a remote device such as a smartphone. The smartphone can process the sensor output and be responsible for warning the subject with an audible output when a threshold (such as glucose concentration) is exceeded. In such an embodiment, the LED and the device network module can be connected to a microprocessor, and the microprocessor monitors the connection state of the module and causes the LED to generate an output when the connection is established and / or when the connection is lost. The application software of the smartphone may be set to warn the subject that the data connection has been lost, but the smartphone may be powered off (e.g., out of charge, etc.), and in that case, the only means to warn the subject is only through the device itself.

[0281] An output function similar to that of the LED may be provided by a buzzer or a small speaker that provides an audible output understandable by the subject. The output may be, for example, a tone, a series of tones, or a synthesized voice.

[0282] Now, refer to another embodiment of the device shown in FIG. 13. This is a modified version of the embodiments shown in FIGS. 8-12. The embodiment of FIG. 13 includes a temperature sensor (900) that extends through the space (905) of the skin contact portion (530) during operation and contacts the surface of the subject's skin. The temperature sensor (900) may be, for example, a thermocouple or a thermistor operably connected to a microprocessor on a PCB (565). The temperature sensor may be in direct contact with the skin or separated from the skin by a thermally conductive material.

[0283] The temperature sensor may be disposed within a pocket or other structure sized to accommodate the temperature sensor. The pocket may be fabricated from a thin sheet-like plastic material such as thermally conductive plastic that includes a metal or other filler to facilitate the transfer of thermal energy from the underlying skin to the temperature sensor. The temperature sensor may be surrounded by a thermally conductive paste to facilitate the transfer of thermal energy from the pocket wall to the temperature sensor.

[0284] The bottom of the pocket may extend outward from the device such that when the device is applied to the skin, the bottom of the pocket is gently pressed against the skin surface, thereby facilitating the transfer of thermal energy from the skin to the temperature sensor. It will be appreciated that pressing the bottom of the pocket too hard against the skin surface can squeeze blood out of the skin's capillaries, thereby artificially cooling the skin surface.

[0285] Preferably, only the bottom of the pocket is fabricated from a thermally conductive material and the remainder is fabricated from a material having a low thermal conductivity. This arrangement prevents thermal energy from escaping from the temperature sensor away from the skin.

[0286] Insulating material can form the ceiling of the pocket to retain thermal energy around the temperature sensor and prevent it from being lost to the internal cavity of the housing.

[0287] The pocket may include a space extending through the bottom such that the temperature sensor can directly contact the skin surface. Given that no thermal energy is required to pass through intervening material, a temperature close to the actual skin temperature is expected.

[0288] In a further modification, the temperature sensor may be an infrared sensor module, in which case at least the material of the pocket bottom should not substantially interfere with its operation. It is contemplated that a space can be formed in the bottom such that the infrared sensor module is directly exposed to the skin surface to accurately read the skin temperature.

[0289] The signal output from the temperature sensor (300) is used in calculations by the microprocessor (or a remote microprocessor) to more accurately determine the concentration of the target analyte. For example, the microprocessor can access the stored calibration curves for which each curve was run at a given temperature. Based on the output of the temperature sensor (300), an appropriate calibration curve is selected and a more accurate analyte concentration is determined.

[0290] The embodiment of FIG. 13 includes a release member (600) having a pair of protrusions (the first protrusion is shown as (650) and the second protrusion of the pair of protrusions is hidden by the first protrusion). The protrusion (650) extends downward through the space (573) of the skin contact portion (530). The function of the protrusion (650) is to prevent the lateral movement of the release member (600) until the lower surface of the skin contact portion (30) is pressed against the skin. When pressed against the skin, the protrusion (650) exits the space (573) vertically and the subject can pull the release member (600) laterally. This mechanism prevents the release member (600) from being accidentally removed before the device is properly applied to the skin surface. Without such a mechanism, the microneedles (515) could extend through the space (545) prematurely and be contaminated by contact with air or objects, or physically damaged, for example, by snagging on clothing.

[0291] In some embodiments of the device, it is necessary to electrically insulate the upper region of the microneedles to avoid the wet surface of the skin (separate from the underlying biological fluid) forming a conductive path between the microneedles.

[0292] As another means of controlling moisture, an absorbent material can be positioned on the micro-needle mount portion and near the tip of the micro-needle. In embodiments of devices for sensing applications, the material is configured to absorb excess liquid that may be generated by inserting the micro-needle into the skin, improving the subject's experience and reducing problems that may arise from liquid contact with other parts of the device, such as electronic circuits or electrical contacts. In embodiments of devices for liquid extraction applications, etc., the material functions as a wicking agent to transport liquid from the micro-needle site to the required final site on or outside the device. In some embodiments, the absorbent material is sheet-like. In embodiments where it is desirable to prevent contamination or damage of the micro-needle prior to insertion, the sheet is provided with holes through which the micro-needle passes, and the dimensions of the holes are large enough to prevent the absorbent material from contacting the micro-needle during the micro-needle insertion process, but small enough for excess liquid exuding from the access penetration point formed by the micro-needle to contact and be absorbed by the material. In other embodiments, such as when the device is intended for use in liquid extraction, the sheet of absorbent material either has no holes or the hole dimensions are set such that the absorbent material contacts the micro-needle during and after insertion to assist its sucking-up action. In embodiments where the sheet has no holes, the micro-needle forms holes as it passes through the sheet as part of the insertion process.

[0293] The device is configured and / or can be used in any suitable application where it is necessary to embed the micro-needle in the subject's skin for an extended period of time.

[0294] Such applications include electrochemical aptamer-based sensing in which a target analyte in a biological fluid is detected by binding to a capture entity such as an aptamer that includes a redox reporter. The capture entity is bound to the micro-needle either covalently or non-covalently, and the redox reporter is arranged such that when the target analyte binds, an electrical signal is transmitted by the micro-needle. The target analyte can be a drug or other exogenous species, or an endogenous species such as a hormone or metabolite.

[0295] When the micro-needle functions as an electrode for detecting an analyte present in a layer of skin, the device can include circuitry and components for electrically exciting the electrode and receiving, measuring, and processing electrical signals resulting from the electrical excitation. According to this embodiment, the micro-needle can include a tip, a shaft, and a base, and an electrical signal is generated by an electrode coated on the surface of the micro-needle or integrated with the micro-needle, transmitted along the shaft of the micro-needle to the base of the micro-needle, and an electrical connection is made to the base or shaft of the micro-needle and the electrical signal is transmitted to an electronic circuit between the electrode. The electrode can be formed near the tip of the micro-needle, at least a part of the shaft of the micro-needle not near the tip of the micro-needle, or both near the tip of the micro-needle and at least a part of the shaft of the micro-needle.

[0296] The micro-needle can be connected to an electronic circuit by various methods known in the art, such as soldering, wire wrapping, or spring-loaded pins. In one embodiment, the micro-needle is attached to pass through a plate or block of dielectric material, and the connection portion of the micro-needle is positioned on or above the surface of the plate or block far from the tip of the micro-needle. Using a zebra strip connection to connect the micro-needle to an electronic circuit can facilitate a robust connection without the need to accurately align the zebra connector to the end of the micro-needle in at least one dimension.

[0297] Another potentially useful application is the delivery of an active substance to the skin. This substance can mainly remain in the skin or potentially enter the systemic circulation. In such applications, the microneedles can be hollow, and the substance is delivered through the lumen of the needle. Alternatively, the microneedles can be coated with the active substance to allow the substance to be released instantaneously into the biological fluid or to be released gradually over a long period. As yet another method, the microneedles themselves can be soluble in the biological fluid and contain the active substance within their bulk so that the active substance is released when the microneedles dissolve. The active substance can be a pharmaceutical composition (e.g., a small molecule, protein, peptide, or nucleic acid) for use as a vaccine, or an immunologically active composition (e.g., an aggregate of proteins).

[0298] A further potential application is to deliver an electric current to the skin for the purpose of muscle stimulation or to stimulate or inhibit a biological process of a subject. Similarly, the device can be used to detect the electric current of the skin of a subject, for example, to detect nerve conduction.

[0299] In any of the above applications, the microneedles can be solid or hollow, as required or desired.

[0300] The length of the microneedles can be selected according to a specific application. Typically, the microneedles need to extend at least under the stratum corneum. The depth of the stratum corneum varies by location. For example, this layer is relatively thick on the sole of the foot and relatively thin on the back of the hand. Therefore, the length of the microneedles extending beyond the housing can be adjusted according to the intended application site.

[0301] In some cases, the microneedles may need to extend well below the stratum corneum, into the epidermis, dermis, and even into the subcutaneous layer including the subcutaneous tissue. Also, the length of the microneedles extending beyond the device can be set accordingly.

[0302] Those skilled in the art will also understand that it may be necessary to set the length of the microneedle according to the intended subject. For example, to contact the subcutaneous tissue of a neonatal subject, generally a relatively short microneedle is required, but to contact an adult subject at the same site, a longer microneedle is needed.

[0303] In some applications, it may be desirable for one microneedle to penetrate deeper into the skin than another microneedle. Thus, the two microneedles may terminate at different distances from the skin surface or may terminate at different distances from the microneedle mount. In some embodiments, the lengths of the two microneedles are different. In other embodiments, the microneedles are the same length and the mount is configured to displace one microneedle axially relative to the other microneedle. For example, the mount may be multi-level where a first electrode extends from a first level and a second electrode extends from a second level.

[0304] In typical applications, the microneedle can extend outward from the device by a distance of about 10 μm to about 5000 μm. In many applications, a distance of about 500 μm to about 4000 μm is useful.

[0305] Those skilled in the art will understand that the invention described herein is capable of further variations and modifications other than those specifically described.

[0306] For example, the movable arm can be moved by the user grasping or pressing a flexible portion of the device housing, by actuating a rotary lever, or by sliding an element along an incline to bias the arm downward.

[0307] The skin contact portion of the device is depicted such that its lower side (skin contact surface) is precisely planar. However, in some embodiments, the skin contact portion may be curved to conform to the surface of a body part such as a finger, wrist, heel, or ear. The skin contact portion may have a certain degree of flexibility (at least in one direction) to conform to the surface of the body part.

[0308] The space through which the microneedles extend is typically shown as an opening, but other types of spaces are also conceivable. In some embodiments, the space is not an opening, and in one such embodiment, it has microneedles that extend through the space around the skin contact portion.

[0309] Those skilled in the art will understand that the invention described herein may be subject to further variations and modifications other than those specifically described. It is understood that the present invention encompasses all such variations and modifications that fall within the spirit and scope of the present invention.

[0310] 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 permitted by law.

Claims

Claim 1 A method for manufacturing an electrochemical aptamer-based sensor device, the method comprising: assembling two or more electrodes of an electrochemical aptamer-based sensor with a device for contacting the two or more electrodes with the skin of a subject; a method. Claim 2 The method according to claim 1, wherein at least one of the two or more electrodes is a working electrode configured to specifically detect an analyte. Claim 3 The method according to claim 1, wherein at least two of the two or more electrodes are each a working electrode configured to specifically detect an analyte. Claim 4 The method according to claim 3, wherein each of the two or more working electrodes contains a different aptamer species, and each of the different aptamer species is configured to specifically detect a different analyte or the same analyte. Claim 5 The method according to any one of claims 1 to 4, wherein the two or more electrodes are assembled with a fixed mutual spacing relationship. Claim 6 The method according to any one of claims 2 to 5, comprising assembling one or more counter electrodes with the working electrodes. Claim 7 The method according to any one of claims 2 to 6, comprising assembling one or more reference electrodes with the working electrodes. Claim 8 The method according to any one of claims 1 to 7, wherein the electrodes are regularly arranged. Claim 9 The method according to claim 8, wherein the regular arrangement is an array. Claim 10 The method according to any one of claims 6 to 9, comprising a counter electrode and two or more working electrodes, each of the two or more working electrodes being substantially equidistant from the counter electrode. Claim 11 The method according to any one of claims 7 to 10, comprising a reference electrode and two or more working electrodes, each of the two or more working electrodes being substantially equidistant from the reference electrode. Claim 12 The method according to any one of claims 1 to 11, wherein all the electrodes are assembled with a fixed mutual spatial relationship. Claim 13 The method according to any one of claims 1 to 12, wherein the distance between any two electrodes is less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, 2 mm, 1 mm, 0.9 mm, 0.8 mm, 0.7 mm, 0.6 mm, 0.5 mm, 0.4 mm, 0.3 mm, 0.2 mm, or 0.1 mm. Claim 14 All electrodes are approximately 100 mm ２ , 90mm ２ , 80 mm ２ , 70 mm ２ , 60 mm ２ , 50 mm ２ , 40 mm ２ , 30 mm ２ , 20mm ２ , 10 mm ２ , 9mm ２ , 8mm ２ , 7mm ２ , 6mm ２ , 5 mm ２ , 4mm ２ , 3 mm ２ , 2 mm ２ , or 1 mm ２ 14. The method of claim 1 , wherein the first and second electrodes are disposed within an area of ​​less than 100 nm. Claim 15 The method according to any one of claims 1 to 14, wherein at least one of said electrodes is a wire, a needle, or a micro-needle.

16. The method according to any one of claims 1 to 15, wherein said assembly includes a step of attaching each electrode to a mounting portion.

17. The method according to claim 16, wherein when the sensor device is assembled, one electrode terminates distally from the other electrode from said mounting portion.

18. The method according to claim 16 or 17, wherein said mounting portion is substantially resistant to bending and / or stretching and / or contraction.

19. The method according to any one of claims 16 to 18, wherein said mounting portion electrically insulates each electrode from each other electrode.

20. The method according to any one of claims 16 to 19, wherein said electrode and / or said mounting portion is configured to form a waterproof seal at a joint formed therebetween.

21. The method according to claim 20, wherein said waterproof seal is formed by press-fitting, snap-fitting, or friction fitting between said electrode and said mounting portion.

22. The method according to claim 20 or 21, wherein said waterproof seal is formed by a flexible seal or a curable sealant applied to or around the joint.

23. The method according to any one of claims 20 to 22, wherein said waterproof seal is formed by a screw connection between said electrode and said mounting portion.

24. The method according to any one of claims 16 to 23, wherein at least one of said electrodes includes an extended region configured to contact the surface of said mounting portion.

25. The method according to any one of claims 1 to 24, including a step of assembling at least 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 electrodes.

26. The method according to any one of claims 1 to 25, wherein each electrode is a wire, a needle, or a micro-needle.

27. The method according to any one of claims 1 to 26, wherein each electrode is obtained by removing an electrode from a group of electrodes having the same analyte specificity, dimensions, material, or function.

28. The method according to claim 27, wherein said group of electrodes is held in a holder configured to removably hold said electrodes.

29. At least one of the electrodes is a working electrode, and the working electrode is selected from an electrode library including at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 working electrodes each containing a different aptamer species, according to any one of claims 1 to 28.

30. The working electrodes containing the same aptamer species are grouped in individual holders or in regions of a single holder, according to the method of claim 29.

31. The device for bringing the two or more electrodes into contact with the skin of the subject is a skin contact portion defining a skin contact surface and one or more spaces through which the two or more electrodes can extend, and a movable part configured to move the two or more electrodes from a first position behind the skin contact surface to a second position protruding from the skin contact surface, according to any one of claims 1 to 30.

32. The device includes a holding part configured to hold the skin contact surface in contact with the skin during use, according to the method of claim 31.

33. The movable part is configured to move from the first position to the second position along a non-linear path, according to the method of claim 31 or 32.

34. The non-linear path is a substantially arc-shaped path, according to the method of claim 33.

35. The movable part has a connection end and a free end, according to any one of claims 31 to 34.

36. The free end moves a longer distance than the connection end, according to the method of claim 35.

37. The non-linear path is drawn with reference to the free end, according to any one of claims 31 to 36.

38. The non-linear path is less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, or 3 mm, according to any one of claims 31 to 37.

39. The angle of the arc is less than about 45°, 40°, 35°, 30°, 25°, 20°, 15°, 14°, 13°, 12°, 11°, 10°, 9°, 8°, 7°, 6°, or 5°, according to any one of claims 34 to 38.

40. The movable part has a swivel part, a hinge part, a bending part, or a mounting part, according to any one of claims 31 to 39.

41. The movable part is associated with the mounting part and is the method according to any one of claims 31 to 40.

42. The method according to claim 41, wherein the mounting part is fixed during use and the movable part is movable relative to the mounting part.

43. The method according to claim 41 or 42, wherein the mounting part includes a portion that allows the movable part to pivot, be hinge-connected, bend, or be attached.

44. The method according to any one of claims 41 to 43, wherein the mounting part is disposed at a certain distance from the skin contact surface.

45. The method according to any one of claims 41 to 44, wherein the mounting part is disposed at a distance of about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, or less than 2 mm from the skin contact surface.

46. The method according to any one of claims 41 to 45, wherein the mounting part is substantially in a lateral direction with respect to the movable part.

47. The device further includes a release part operable by the user, and the release part is configured to hold the movable part in the first position until the user operates the release part. When the user operates the release part, the movable part is released and can move to the second position. The method according to any one of claims 31 to 46.

48. The device further includes a locking part configured to lock the movable part when the movable part is in the second position. The method according to any one of claims 31 to 47.

49. The device is configured such that a motive power generated inside and / or outside the device is required to move the movable part from the first position to the second position. The method according to any one of claims 31 to 48.

50. The motive power inside the device is generated from a spring, an elastically deformable member, a shape memory member, or other biasing means, and the motive power outside the device is generated from the user. The method according to claim 49.

51. The device does not have an internal motive power generator configured to move the movable part from the first position to the second position. The method according to any one of claims 31 to 50.

52. The method according to any one of claims 31 to 51, wherein the holding part is a dermatologically acceptable composition disposed on or around the skin contact surface, or contains this composition.

53. The method according to claim 52, wherein the dermatologically acceptable composition is an adhesive or a functional equivalent thereof.

54. The method according to any one of claims 31 to 53, wherein the holding part is configured to mechanically hold the skin contact surface in contact with the skin.

55. The method according to claim 54, wherein the holding part is selected from one or more of a strap, a band, a belt, a clamp, a grip, a necktie, a fastener, a sleeve, a stocking, a sock, a glove, a cap, a hat, pants, a singlet, a shirt, a bra, a top, trousers, a scarf, a ring, glasses, and a choker.

56. The method according to any one of claims 31 to 55, wherein the two or more electrodes are mechanically connected directly or indirectly to the movable part.

57. The method according to any one of claims 31 to 56, wherein the two or more electrodes are wires, needles, and / or microneedles.

58. The method according to claim 57, wherein the two or more electrodes form an array.

59. The method according to any one of claims 31 to 58, wherein the two or more electrodes are long enough to contact the epidermis, dermis, or subcutaneous tissue of the subject.

60. The two or more electrodes are configured to conduct an electric current to, from, or through the skin, conduct a sound wave to, from, or through the skin, conduct light to, from, or through the skin, conduct heat to, from, or through the skin, sample a body fluid or tissue from the skin, deliver a biologically active substance to the skin, or introduce an analyte sensing substance into the skin during use, according to the method of any one of claims 31 to 59.

61. The two or more electrodes are each conductive, and the device further includes a circuit having an audio, visual, or tactile indicator, the circuit being configured to activate the indicator when the one or more protrusions contact a conductive fluid that naturally exists on the skin, the method according to any one of claims 31 to 60.

62. The circuit includes at least two protrusions, and the circuit is configured to be completed by activating the indicator when the at least two protrusions contact the conductive fluid that naturally exists on the skin, the method according to claim 61.

63. The circuit includes one protrusion and at least one conductive pad disposed in contact with the skin, the circuit being configured to be completed by electrically communicating the protrusion and the pad with the conductive fluid that naturally exists on the skin to activate the indicator, the method according to claim 61.

64. The device includes a housing dimensioned such that when the device is applied to the skin, the movable part is in the second position, and any part of each of the two or more electrodes protruding from the skin contact surface is embedded in the skin, the housing extends at or below about 5 mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 11 mm, 12 mm, 13 mm, 14 mm, 15 mm, 16 mm, 17 mm, 18 mm, 19 mm, or 20 mm over most or substantially all of the skin, the method according to any one of claims 31 to 63.

65. The device is configured to be used for a period exceeding about 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 24 hours, 36 hours, 48 hours, 60 hours, 72 hours, 84 hours, or 96 hours, the method according to claim 31 or 32.

66. The device is configured such that the two or more electrodes are inseparable from the device or cannot be separated without the assistance of tools, the method according to any one of claims 31 to 65.

67. The movable part and the mount part are integrated, the method according to any one of claims 41 to 66.

68. The integrated movable part and mount part are manufactured from an elastically deformable material, the method according to claim 67.

69. The integrated movable part and mount part are part of a circuit board of the device, according to the method of claim 35 or 36.

70. The movable part is biased toward the second position and is maintained in the first position and against the bias by a release part operable by the user until the release part operates. When the release part operates, the movable part is released and can move to the second position, according to the method of any one of claims 47 to 69.

71. The release part operable by the user is a shelf configured to hold the movable part in the first position, and by the motive force provided by the user deforming the shelf and / or the movable part, the movable part is released from the shelf and can move to the second position, according to the method of any one of claims 47 to 69.

72. The movable part is hingedly connected in relation to the skin contact part, according to the method of any one of claims 31 to 71.

73. The hinge is arranged in or towards a peripheral area of the movable part and the skin contact part, according to the method of claim 72.

74. The release part includes a member configured to maintain the movable part in the first position, but is removable or deformable by the user so that the movable part can move to the second position, according to the method of any one of claims 47 to 73.

75. The member is removable by sliding substantially across the skin contact part, according to the method of claim 74.

76. The member is substantially wedge-shaped, the device includes a hinge associating the movable part with the skin contact part, the thin part of the wedge is arranged proximal to the hinge, and the thick part of the wedge is arranged distal to the hinge, according to the method of claim 74 or 75.

77. The release part is removable from the device and includes a gripping part for facilitating manual removal, according to the method of any one of claims 74 to 76.

78. An electrochemical aptamer-based sensor device, the device comprising an assembly of two or more electrodes and an assembly of a device for bringing the two or more electrodes into contact with the skin of a subject. Device.

79. The device according to claim 78, wherein at least one of the two or more electrodes is a working electrode including an aptamer species configured to specifically detect an analyte.

80. The device according to claim 78, wherein at least two of the two or more electrodes are working electrodes each including a different aptamer species.

81. The device according to any one of claims 78 to 80, wherein the two or more electrodes are assembled in a fixed mutual spatial relationship.

82. The device according to any one of claims 78 to 81, wherein one of the two or more electrodes is a counter electrode.

83. The device according to any one of claims 78 to 81, wherein one of the two or more electrodes is a reference electrode.

84. The device according to claim 83, wherein the electrodes are regularly arranged.

85. The device according to claim 84, wherein the regular arrangement is an array.

86. The device according to any one of claims 78 to 85, wherein the two or more electrodes include a counter electrode and two or more working electrodes, and each of the two or more working electrodes is substantially equidistant from the counter electrode.

87. The device according to any one of claims 78 to 86, wherein the two or more electrodes include a reference electrode and two or more working electrodes, and each of the two or more working electrodes is substantially equidistant from the reference electrode.

88. The device according to any one of claims 78 to 87, wherein all the electrodes are arranged in a fixed mutual spatial relationship.

89. The device according to any one of claims 78 to 88, wherein the distance between any two electrodes is less than about 10 mm, 9 mm, 8 mm, 7 mm, 6 mm, 5 mm, 4 mm, 3 mm, 2 mm, 1 mm, 0.9 mm, 0.8 mm, 0.7 mm, 0.6 mm, 0.5 mm, 0.4 mm, 0.3 mm, 0.2 mm, or 0.1 mm.

90. All electrodes are approximately 100 mm ２ , 90mm ２ , 80 mm ２ , 70 mm ２ , 60 mm ２ , 50 mm ２ , 40 mm ２ , 30 mm ２ , 20mm ２ , 10 mm ２ , 9mm ２ , 8mm ２ , 7mm ２ , 6mm ２ , 5 mm ２ , 4mm ２ , 3 mm ２ , 2 mm ２ , or 1 mm ２ 90. The device of any one of claims 78 to 89, wherein the device is positioned within an area of ​​less than

91. The device according to any one of claims 78 to 90, wherein at least one electrode is a wire, a needle, or a micro-needle.

92. The device according to any one of claims 78 to 91, wherein the at least two electrodes are attached to a mounting portion.

93. The device according to claim 92, wherein the mounting portion is substantially resistant to bending and / or stretching and / or contraction.

94. The device according to claim 92 or 93, wherein the mounting part electrically insulates each electrode from the other electrodes.

95. The device according to any one of claims 92 to 94, wherein the electrodes are formed separately from the mounting part, and the electrodes and the mounting part are assembled to form the device.

96. The device according to claim 95, wherein the electrodes and / or the mounting part are configured to form a waterproof seal at a joint formed therebetween.

97. The device according to claim 96, wherein the waterproof seal is formed by press-fitting, snap-fitting, or friction fitting between the electrode and the mounting part.

98. The device according to claim 96 or 97, wherein the waterproof seal is formed by a flexible seal or a curable sealant applied to or around the joint.

99. The device according to any one of claims 96 to 98, wherein the waterproof seal is formed by a screw connection between the electrode and the mounting part.

100. The device according to any one of claims 92 to 99, wherein at least one of the two or more electrodes includes an extended region configured to contact the surface of the mounting part.

101. The device according to any one of claims 78 to 100, including at least 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 electrodes.

102. The device according to any one of claims 78 to 101, wherein each electrode is a wire, a needle, or a micro-needle.

103. The device according to any one of claims 78 to 102, wherein each electrode is obtained by removing one electrode from a group of electrodes having the same analyte specificity, dimension, material, or function.

104. The device according to claim 103, wherein the group of electrodes is held in a holder configured to removably hold the electrodes.

105. The device according to any one of claims 78 to 104, wherein at least one of the electrodes is a working electrode, and the working electrode is selected from an electrode library including at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 working electrodes having different analyte specificities.

106. The device according to claim 105, wherein the electrodes of a given analyte specificity are grouped in individual holders or in regions of a single holder.

107. The device according to any one of claims 78 to 106, having the characteristics of the device defined in any one of claims 31 to 77.

108. A system for manufacturing an electrochemical aptamer-based sensor device, the system comprising: a library of two or more electrodes of the electrochemical aptamer-based sensor device; and a mount configured to attach the two or more electrodes in a mutually spaced relationship; wherein the mount is provided by a device having the characteristics defined in any one of claims 31 to 77. System.

109. The system according to claim 108, wherein the two or more electrodes are each working electrodes, and each working electrode contains a different aptamer species.

110. The system according to claim 108 or 109, wherein the electrode library includes at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 working electrodes containing different aptamer species.

111. The system according to claim 110, wherein the working electrodes containing the same aptamer species are grouped in individual holders or in regions of a single holder.

Images