CA3171868A1

CA3171868A1 - Devices, systems, and methods for measuring a solution characteristic of a sample using a multi-layered active sensor

Info

Publication number: CA3171868A1
Application number: CA3171868A
Authority: CA
Inventors: Creighton T. BUIE; Nitin K. RAJAN; Andrew H. Theiss; Elizabeth Botbol PONTE; Meike Herget; Oren S. KNOPFMACHER; Michael D. Laufer
Original assignee: Avails Medical Inc
Current assignee: Avails Medical Inc
Priority date: 2020-05-15
Filing date: 2021-05-13
Publication date: 2021-11-18
Also published as: EP4150321A2; WO2021231718A2; WO2021231718A3; JP2023525197A; CN115004010A

Abstract

Various apparatus, systems, and methods for measuring a solution characteristic of a sample comprising microorganisms are disclosed. In one embodiment, a sensor apparatus is disclosed comprising a sample chamber having a chamber lateral wall surrounding a chamber cavity configured receive the sample, a reference sensor comprising a wicking component to wick the sample to a reference electrode material, and an active sensor made of a substrate covered in part by an active electrode layer. The active sensor can be coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall. The active sensor can be positioned such that the active electrode layer faces the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

Description

DEVICES, SYSTEMS, AND METHODS FOR MEASURING A SOLUTION
CHARACTERISTIC OF A SAMPLE USING A MULTI-LAYERED ACTIVE
SENSOR
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application claims the benefit of U.S. Provisional Application No.
63/025,575 filed on May 15, 2020, the content of which is incorporated herein by reference in its entirety.
TECHNICAL FIELD

[0002] The present disclosure relates generally to diagnostic devices for measuring a solution characteristic of a sample; more specifically, to devices, systems, and methods for measuring a solution characteristic of a sample using a multi-layered active sensor.
BACKGROUND

[0003] Infections caused by anti-infective resistant infectious agents or microbes are a significant problem for healthcare professionals in hospitals, nursing homes, and other healthcare environments. For example, such infections can lead to a potentially life-threatening complication known as sepsis where chemicals released into the bloodstream by an infectious agent can trigger a dangerous whole-body inflammatory response as well as a vasoactive response causing fever, low blood pressure, and possibly death. When faced with such an infection, a preferred course of action is for a clinician to use anti-infective compounds judiciously, preferably only those necessary to alleviate the infection.

[0004] However, what occurs most frequently today is that until the organism is identified and tested for drug sensitivity, broad-spectrum anti-infectives, often multiple drugs, are given to the patient to ensure adequacy of treatment. This tends to result in multiple drug-resistant infectious agents. Ideally, the sensitivity of the infectious agent would be detected soon after its presence is identified. In order to determine the susceptibility of such infectious agents to anti-infectives, samples comprising such infectious agents must be quantified, which requires such samples to be assayed for microbial growth or lack thereof.

[0005] Existing biosensors used to assay infectious agents and pathogens in biological or other types of samples often include an active sensing component and a reference sensing component that are in fluid contact or communication with the sample of interest.
Current in vitro diagnostic measurement systems, especially those used to detect oxidation reduction potentials (ORP) and pH in biological or fluid samples, are often not designed with both high-performance and low-cost considerations in mind. Moreover, since it is important to prevent cross-contamination of patient samples, a single-use disposable consumable is the preferred design for the sensing component of the diagnostic measurement system. This puts significant emphasis on the cost and manufacturability of a single-use disposable sensing component.

[0006] Traditional biosensors are often made using costly glass or silicon substrates that drive up the cost of such sensors and require numerous manufacturing steps to produce. Moreover, the active sensing component of such biosensors may malfunction when the biological sample or other fluid sample makes inadvertent contact with the conductive parts of the active sensing not intended to contact the sample.

[0007] Therefore, a solution is needed that addresses the above shortcomings and limitations. Such a solution should be single-use and cost-effective to manufacture.
However, such a solution should also provide accurate measurements and be made of biocompatible materials.
SUMMARY

[0008] Disclosed are apparatus, systems, and methods for measuring a solution characteristic (e.g., an ORP or a pH) of a sample comprising microorganisms.
In one embodiment, a sensor apparatus for measuring a solution characteristic of a sample is disclosed. The sensor apparatus can comprise a sample container comprising a sample chamber. The sample chamber can comprise a chamber lateral wall surrounding a chamber cavity configured to receive the sample. The sensor apparatus can also comprise a reference sensor comprising a reference electrode material and a wick in fluid communication with the sample chamber. At least some of the sample can be drawn up by the wick in a direction of the reference electrode material.

[0009] The sensor apparatus can also comprise an active sensor made of a conductive substrate covered in part by an active electrode layer. The active sensor can be coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall. In some embodiments, no part of the active sensor extends into the chamber cavity. The active electrode layer can face the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening. The solution characteristic of the sample can be determined based on a potential difference measured between the active sensor and the reference sensor when the reference sensor and the active sensor are electrically coupled to a reader apparatus.

[0010] The solution characteristic measured can be an oxidation reduction potential.
The solution characteristic measured can be pH. The active electrode layer can comprise at least one of a platinum oxide layer (formed on a platinum layer) and a tantalum oxide layer. The conductive substrate can be stainless steel. The active electrode layer can have an active electrode layer thickness of between about 50 nm and 500 nm. The active sensor can further comprise an adhesion layer between the conductive substrate and the active electrode layer. The adhesion layer can have a thickness of between 5 nm and 50 nm. A
ratio of the adhesion layer thickness to the active electrode layer thickness can be between about 1:10 and 1:20. The adhesion layer can be a chromium layer, a gold layer, or a nickel layer.

[0011] The active layer can have an active electrode layer thickness of 400 nm. The active sensor can be insert-molded into the chamber lateral wall while the sample container is formed by injection molding. The active sensor can be press-molded into the chamber lateral wall after the sample container is formed by injection molding. The chamber lateral wall can comprise a recessed portion surrounding the window opening. The recessed portion can be defined along an exterior side of the chamber lateral wall. The active sensor can be adhered to at least part of the chamber lateral wall within the recessed portion via an adhesive.

[0012] The active sensor can comprise an active electrode side, a conductive substrate side opposite the active electrode side, and lateral sides. The lateral sides can be covered by at least one of the chamber lateral wall and an adhesive to prevent the lateral sides from contacting the sample. The sample chamber can be made in part of at least one of polyoxymethylene, polyamide, polyethylene, acrylonitrile butadiene styrene, polycarbonate, and polypropylene. The reference electrode material can be a cured or hardened electrically-conductive ink deposited or otherwise applied on a wick proximal end of the wick.

[0013] In some embodiments, the active sensor can be made of a non-conductive printed circuit board (PCB) substrate covered in part by an electrode layer.
The active electrode layer can be electrically coupled to conductive contacts of the PCB
substrate by a conductive via extending through the PCB substrate. The PCB substrate can be a flexible PCB substrate.

[0014] The PCB substrate can be made in part of polyimide, an FR-4 composite material, copper, or the like. The solution characteristic measured can be an oxidation reduction potential. The active electrode layer can be a platinum layer or a gold layer. The active electrode layer can have an active electrode layer thickness of at least 50 nm. In certain embodiments, the active electrode layer can have an active electrode layer thickness of at least 400 nm.

[0015] In some embodiments, the active sensor can be made of a non-conductive polymeric substrate comprising a through-hole. One side of the polymeric substrate and one end of the through-hole can be covered by a conductive layer. The active electrode layer can be electrically coupled to the conductive layer via a conductive coating covering lateral sides of the through-hole.

[0016] The active electrode layer can be a platinum layer or a gold layer.
The active electrode layer can have an active electrode layer thickness of at least 50 nm. In certain embodiments, the active electrode layer can have an active electrode layer thickness of at least 400 nm.

[0017] The active sensor can be a rectangular piece having a width dimension of between about 100 um and 6.0 mm and a length dimension of between about 100 um and 6.0 mm. The through-hole can have a diameter between about 10 um to 100 um.
The conductive coating covering the lateral sides of the through-hole can be a coating of platinum, gold, or the like.

[0018] In some embodiments, the active sensor can be made of a conductive dowel covered in part by an active electrode layer. The active sensor can be coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall. The part of the conductive dowel covered by the active electrode layer can extend into the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer. An end of the conductive dowel not extending into the chamber cavity can extend out of the chamber lateral wall. The conductive dowel can be made in part of stainless steel and can be shaped substantially as a cylinder having rounded edges.

[0019] Also disclosed is a method of measuring a solution characteristic of a sample.
The method can comprise cleaning a conductive substrate with an acid and base treatment, depositing an adhesion layer on one side of the conductive substrate, and depositing an active electrode layer on the adhesion layer. The method can also comprise singulating the conductive substrate covered by the adhesion layer and the active electrode layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber. The method can also comprise coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

[0020] The method can also comprise treating the conductive substrate with nitric acid followed by treating the conductive substrate with ammonium hydroxide, isopropyl alcohol, or acetone. The method can also comprise laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or sawing the conductive substrate.

[0021] The method can also comprise applying a bead of adhesive to a part of the chamber lateral wall within a recessed portion defined along the chamber lateral wall surrounding the window opening. The method can also comprise pressing the active sensor onto the bead of adhesive within the recessed portion and curing the adhesive.

[0022] The method can also comprise insert-molding the active sensor into the chamber lateral wall while the sample chamber is formed by injection molding.
The method can also comprise focally melting a part of the chamber lateral wall surrounding the window opening, pressing the active sensor onto the melted part of the chamber lateral wall, and allowing the melted part of the chamber lateral wall to cool to affix the active sensor to the chamber lateral wall.

[0023] The method can also comprise depositing an active electrode material making up the active electrode layer until a thickness of the active electrode layer is at least 50 nm.
In some embodiments, the method can comprise depositing an active electrode material making up the active electrode layer until a thickness of the active electrode layer is at least 400 nm. In some embodiments, the active electrode material can be platinum when the solution characteristic measured is an oxidation reduction potential (ORP) of the sample.
The active electrode material can be deposited using sputter deposition, evaporation deposition, electrodeposition, or ink screen-printing.

[0024] The method can also comprise depositing an adhesion material making up the adhesion layer using sputter deposition. The adhesion layer can be deposited in a vacuum chamber and the active electrode layer can be deposited subsequent to the adhesion layer in the same vacuum chamber.

[0025] The active electrode material can be a metal oxide when the solution characteristic measured is a pH of the sample. In some embodiments, the metal oxide can be platinum oxide and the platinum oxide can cover a platinum layer deposited on the adhesion layer.

[0026] In some embodiments, a method of making a sensor apparatus for measuring a solution characteristic of a sample can comprise providing a non-conductive printed circuit board (PCB) substrate and depositing an active electrode layer on one side of the PCB
substrate. The active electrode layer, after the deposition step, can be electrically coupled to conductive contacts of the PCB substrate by conductive vias extending through the PCB
substrate. The method can also comprise singulating the PCB substrate covered by the active electrode layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber. The active sensor can comprise at least one conductive via extending through the PCB substrate. The method can also comprise coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

[0027] The method can also comprise depositing an active electrode material making up the active electrode layer using sputter deposition, evaporation deposition, and electrodeposition. An active electrode material can be deposited until a thickness of the active electrode layer is at least 50 nm. In certain embodiments, the active electrode material can be deposited until a thickness of the active electrode layer is at least 400 nm.
The active electrode material can be platinum or gold when the solution characteristic measured is an oxidation reduction potential (ORP) of the sample. The conductive contacts can be made in part of gold.

[0028] In some embodiments, another method of making a sensor apparatus can comprise providing a non-conductive polymeric substrate comprising a plurality of through-holes and depositing a conductive layer on one side of the polymeric substrate.
The method can also comprise depositing an active electrode layer on the other side of the polymeric substrate. One end of each of the through-holes can be covered by the active electrode layer and the other end of each of the through-holes can be covered by the conductive layer. The active electrode layer, after the deposition steps, can be electrically coupled to the conductive layer via a conductive coating covering lateral sides of the through-holes. The method can also comprise singulating the polymeric substrate covered by the active electrode layer and the conductive layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber. The active sensor can comprise at least one through-hole covered by the active electrode layer and the conductive layer.

[0029] The method can also comprise coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.
Depositing the conductive layer can comprise depositing a conductive material on the other side of the polymeric substrate. In some embodiments, the conductive material can be gold.
BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Fig. 1A illustrates a front view of one embodiment of a sensor apparatus for measuring a solution characteristic of a sample.

[0031] Fig. 1B illustrates a cross-sectional side view of part of the sensor apparatus.

[0032] Fig. 1C illustrates a perspective close-up view of an active sensor of the sensor apparatus adhered to a chamber lateral wall of the sensor apparatus.

[0033] Fig. 1D illustrates a sectional view of a sample-filled sensor apparatus.

[0034] Fig. 2 illustrates an embodiment of an active sensor of the sensor apparatus insert molded into a chamber lateral wall of the sensor apparatus.

[0035] Fig. 3A is a black-and-white image of a top plan view of a side of the active sensor covered by an active electrode layer. The active sensor is molded into part of the chamber lateral wall in this image.

[0036] Fig. 3B is a black-and-white image of the opposite side of the active sensor shown in Fig. 3A. The active sensor is molded into part of the chamber lateral wall in this image.

[0037] Fig. 4A illustrates a perspective view of one embodiment of an active sensor.

[0038] Fig. 4B illustrates a side view of one embodiment of an active sensor used for measuring ORP.

[0039] Fig. 4C illustrates a side view of another embodiment of an active sensor used for measuring pH.

[0040] Fig. 4D illustrates a side view of another embodiment of an active sensor used for measuring pH.

[0041] Fig. 5A illustrates a side view of another embodiment of an active sensor made using a PCB substrate.

[0042] Fig. 5B illustrates a single PCB board covered by an active electrode layer that can be singulated into numerous individual active sensors.

[0043] Fig. 6A is a black-and-white image showing an active sensor comprising three individual active electrodes.

[0044] Fig. 6B is a black-and-white image showing a close-up view of a contact side of the active sensor shown in Fig. 6A.

[0045] Fig. 6C is a black-and-white image showing a single PCB board that can be singulated into numerous individual active sensors.

[0046] Fig. 7 illustrates yet another embodiment of an active sensor made by covering a non-conductive polymeric substrate comprising a through-hole with an active electrode layer and a conductive contact layer.

[0047] Figs. 8A and 8B are side cross-sectional views illustrating two different embodiments of the active sensor.

[0048] Fig. 9 illustrates that a large sheet of non-conductive plastic or a large PCB can be singulated into numerous active sensors.

[0049] Fig. 10 illustrates a side view of another embodiment of a sensor apparatus comprising an active sensor made of a conductive dowel.

[0050] Fig. 11 is a graph illustrating a change in the oxidation reduction potential (ORP) of three samples containing E. coli measured over time using three different sensors.

[0051] Fig. 12 is a graph illustrating a change in the pH of four samples containing different starting concentrations of E. coli measured over time using the sensor apparatus disclosed herein.

[0052] Fig. 13A illustrates a perspective view of a reader apparatus designed to receive the sensor apparatus and determine a solution characteristic of a sample within the sensor apparatus.

[0053] Fig. 13B illustrates a partial cutaway view of the reader apparatus with a sample-filled sensor apparatus positioned within the reader apparatus.

[0054] Fig. 13C illustrates a perspective view of a portion of the reader apparatus with the reader housing removed.

[0055] Fig. 13D illustrates a close-up view of a gas nozzle of the reader apparatus being connected to the bottom of the sensor apparatus to aerate the sample within the sensor apparatus.

[0056] Fig. 14 illustrates one embodiment of a method of making a sensor apparatus for measuring a solution characteristic of a sample.

[0057] Fig. 15 illustrates another embodiment of a method of making a sensor apparatus for measuring a solution characteristic of a sample.

[0058] Fig. 16 illustrates yet another embodiment of a method of making a sensor apparatus for measuring a solution characteristic of a sample.
DETAILED DESCRIPTION

[0059] Variations of the devices, systems, and methods described herein are best understood from the detailed description when read in conjunction with the accompanying drawings. It is emphasized that, according to common practice, the various features of the drawings may not be to scale. The dimensions of certain features have been expanded or reduced for clarity and not all features may be visible or labeled in every drawing. The drawings are taken for illustrative purposes only and are not intended to define or limit the scope of the claims to that which is shown.

[0060] Figs. 1A-1D illustrate embodiments of a sensor apparatus 100 for measuring a solution characteristic of a sample. In some embodiments, the solution characteristic measured can be an oxidation reduction potential (ORP) of the sample. In other embodiments, the solution characteristic measured can be a pH of the sample.

[0061] In some embodiments, the sample can be obtained from a patient or subject. In other embodiments, the sample can be a biological sample, an environmental sample, or a food sample.

[0062] When the sample is obtained from a patient or subject, the sample can comprise at least one of a bodily fluid of the patient or subject and a swab obtained from the patient or subject.

[0063] In some embodiments, the patient or subject can be a human patient or subject.
In other embodiments, the patient or subject can be a non-human animal patient or subject.

[0064] In some embodiments, the bodily fluid can comprise blood, urine, serum, plasma, saliva, sputum, semen, breast milk, joint fluid, spinal fluid such as cerebrospinal fluid, wound material, mucus, fluid accompanying stool, vaginal secretions, synovial fluid, pleural fluid, peritoneal fluid, pericardial fluid, amniotic fluid, or a combination thereof.

[0065] In these and other embodiments, the swab obtained from the patient or subject can comprise a wound swab, a rectal swab, a vaginal swab, re-suspended instances of the aforementioned swabs, or a combination thereof.

[0066] In all such embodiments, the sample can comprise a number of microorganisms or infectious agents. The apparatus, systems, and methods disclosed herein can be used to assay the sample for microbial growth or lack thereof as part of a microbial quantification procedure or an antibiotic susceptibility testing (AST) procedure.

[0067] In certain embodiments, the sample can comprise or refer to a bacterial culture derived from at least one of a sample obtained from a patient or subject, a biological sample, an environmental sample, and a food sample. For example, the sample can comprise or refer to a bacterial culture or a re-suspended bacterial culture derived from a bodily fluid or swab obtained from a patient or subject. As a more specific example, the sample can comprise a bacterial culture or a re-suspended bacterial culture derived from a bodily fluid or swab obtained from a patient or subject that has tested positive for microorganism growth.

[0068] More specifically, the sample can comprise a bacterial culture derived from blood obtained from a patient or subject that has tested positive for microorganism growth.
In some embodiments, the sample can be or refer to a positive blood culture.
For purposes of this disclosure, a positive blood culture can be a bacterial culture derived from blood drawn from a patient or subject that has tested positive for bacterial growth.
For example, a patient can show symptoms of sepsis (e.g., high fever, chills, etc.) and blood (e.g., 5 mL to mL) can be drawn from the patient and transferred into a commercial blood culturing container or vessel that contain bacterial growth media (e.g., 30 mL to 40 mL
of growth media). The blood culturing container or vessel can then be incubated at 35 C
2 C to allow the bacteria to proliferate. If the patient's blood is contaminated with bacteria, the bacteria will replicate within the container or vessel. A blood culturing system or apparatus can then be used to monitor for bacterial growth (such as by monitoring bacterial CO2 production within the container or vessel) and the system or apparatus can determine the sample as testing "positive" for bacterial growth when a critical CO2 threshold has been met. Depending on the pathogen type and growth rate, the blood culture can turn positive between 7 hours and 3 days. Such a "positive blood culture" can then be used for further downstream testing such as using any of the apparatus, systems, and methods disclosed herein.

[0069] In additional embodiments, the sample can comprise an environmental sample obtained from a stream, river, lake, ocean, contamination site, quarantine zone, an emergency area, or a combination thereof. In other embodiments, the sample can comprise a food sample obtained from a food preparation facility, a dining establishment, a waste facility, or a combination thereof.

[0070] In some embodiments, an aqueous growth media can be added to the sample prior to being introduced into a sample container 104 of the sensor apparatus 100. In other embodiments, the aqueous growth media can be added to the sample once the sample has been injected, delivered, poured, or otherwise introduced into the sample container 104.

[0071] In one embodiment, the aqueous growth media can be a glucose supplemented Mueller Hinton broth (MHG). In other embodiments, the aqueous growth media can be a solution containing bacto-tryptone, tryptic soy digest, yeast extract, beef extract, cation-adjusted Mueller Hinton Broth (CAMHB), starch, acid hydrolysate of casein, calcium chloride, magnesium chloride, sodium chloride, blood or lysed blood including lysed horse blood (LHB), CAMHB-LHB, glucose or other carbohydrates, or a combination thereof.

[0072] The microorganisms or infectious agents that can be assayed using the apparatus, methods, and systems disclosed herein can be any metabolizing single- or multi-cellular organism including bacteria and fungi. In certain embodiments, the microorganisms or infectious agents can be bacteria including, but not limited to, Acinetobacter, Acetobacter, Actinomyces, Aerococcus, Aeromonas, Agrobacterium, Anaplasma, Azorhizobium, Azotobacter, Bacillus, Bacteriodes, Bartonella, Bordetella, Borrelia, Brucella, Burkholderia, Calymmatobacterium, Campylobacter, Chlamydia, Chlamydophila, Citrobacter, Clostridium, Corynebacterium, Coxiella, Ehrlichia, Enterobacter, Enterococcus, Escherichia, Francisella, Fusobacterium, Gardnerella, Haemophilus, Helicobacter, Klebsiella, Lactobacillus, Legionella, Listeria, Methanobacterium, Microbacterium, Micrococcus, Morganella, Moraxella, Mycobacterium, Mycoplasma, Neisseria, Pandoraea, Pasteurella, Peptostreptococcus, Porphyromonas, Prevotella, Proteus, Providencia, Pseudomonas, Ralstonia, Raoultella, Rhizobium, Rickettsia, Rochalimaea, Rothia, Salmonella, Serratia, Shewanella, Shigella, Spirillum, Staphylococcus, Strenotrophomonas, Streptococcus, Streptomyces, Treponema, Vibrio, Wolbachia, Yersinia, or a combination thereof. In other embodiments, the microorganisms or infectious agents can be one or more fungi selected from the genera Candida or Cryptococcus or mold.

[0073] Other specific bacteria that can be assayed using the methods and systems disclosed herein can comprise Staphylococcus aureus, Staphylococcus lugdunensis, coagulase-negative Staphylococcus species (including but not limited to Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus capitis, not differentiated), Enterococcus faecalis, Enterococcus faecium (including but not limited to Enterococcus faecium and other Enterococcus spp., not differentiated, excluding Enterococcus faecalis), Streptococcus pneumoniae, Streptococcus pyo genes, Streptococcus agalactiae, Streptococcus spp., (including but not limited to Streptococcus mitis, Streptococcus pyogenes, Streptococcus gallolyticus, Streptococcus agalactiae, Streptococcus pneumoniae, not differentiated), Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella spp. (including but not limited to Klebsiella pneumoniae, Klebsiella oxytoca, not differentiated), Escherichia coli, Enterobacter spp. (including but not limited to Enterobacter cloacae, Enterobacter aerogenes, not differentiated), Proteus spp.
(including but not limited to Proteus mirabilis, Proteus vulgaris, not differentiated), Citrobacter spp. (including but not limited to Citrobacter freundii, Citrobacter koseri, not differentiated), Serratia marcescens, Candida albicans, Candida glabrata, and Candida tropicalis.

[0074] Other more specific bacteria that can be assayed can comprise Acinetobacter baumannii, Actinobacillus spp., Actinomycetes, Actinomyces spp. (including but not limited to Actinomyces israelii and Actinomyces naeslundii), Aeromonas spp. (including but not limited to Aeromonas hydrophila, Aeromonas veronii biovar sobria (Aeromonas sobria), and Aeromonas caviae), Anaplasma phagocytophilum, Alcaligenes xylosoxidans, Actinobacillus actinomycetemcomitans, Bacillus spp. (including but not limited to Bacillus anthracis, Bacillus cereus, Bacillus subtilis, Bacillus thuringiensis, and Bacillus stearothermophilus), Bacteroides spp. (including but not limited to Bacteroides fragilis), Bartonella spp. (including but not limited to Bartonella bacilliformis and Bartonella henselae, Bifidobacterium spp., Bordetella spp. (including but not limited to Bordetella pertussis, Bordetella parapertussis, and Bordetella bronchiseptica), Borrelia spp.
(including but not limited to Borrelia recurrentis, and Borrelia burgdorferi), Brucella spp.
(including but not limited to Brucella abortus, Brucella canis, Brucella melintensis and Brucella suis), Burkholderia spp. (including but not limited to Burkholderia pseudomallei and Burkholderia cepacia), Campylobacter spp. (including but not limited to Campylobacter jejuni, Campylobacter coli, Campylobacter lari and Campylobacter fetus), Capnocytophaga spp., Cardiobacterium hominis, Chlamydia trachomatis, Chlamydophila pneumoniae, Chlamydophila psittaci, Citrobacter spp., Coxiella bumetii, Corynebacterium spp. (including but not limited to, Corynebacterium diphtheriae, Corynebacterium jeikeum and Corynebacterium), Clostridium spp. (including but not limited to Clostridium perfringens, Clostridium difficile, Clostridium botulinum and Clostridium tetani), Eikenella corrodens, Enterobacter spp. (including but not limited to Enterobacter aerogenes, Enterobacter agglomerans, Enterobacter cloacae and Escherichia coli, including opportunistic Escherichia coli, including but not limited to enterotoxigenic E. coli, enteroinvasive E. coli, enteropathogenic E. coli, enterohemorrhagic E. coli, enteroaggregative E. coli and uropathogenic E. coli), Enterococcus spp.
(including but not limited to Enterococcus faecalis and Enterococcus faecium), Ehrlichia spp.
(including but not limited to Ehrlichia chafeensia and Ehrlichia canis), Erysipelothrix rhusiopathiae, Eubacterium spp., Francisella tularensis, Fusobacterium nucleatum, Gardnerella vaginalis, Gemella morbillorum, Haemophilus spp. (including but not limited to Haemophilus influenzae, Haemophilus ducreyi, Haemophilus aegyptius, Haemophilus parainfluenzae, Haemophilus haemolyticus and Haemophilus parahaemolyticus), Helicobacter spp. (including but not limited to Helicobacter pylori, Helicobacter cinaedi and Helicobacter fennelliae), Kingella kingii, Klebsiella spp. (including but not limited to Klebsiella pneumoniae, Klebsiella granulomatis and Klebsiella oxytoca), Lactobacillus spp., Listeria monocytogenes, Leptospira interrogans, Legionella pneumophila, Leptospira interrogans, Peptostreptococcus spp., Moraxella catarrhalis, Morganella spp., Mobiluncus spp., Micrococcus spp., Mycobacterium spp. (including but not limited to Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium intracellulare, Mycobacterium avium, Mycobacterium bovis, and Mycobacterium marinum), Mycoplasm spp. (including but not limited to Mycoplasma pneumoniae, Mycoplasma hominis, and Mycoplasma genitalium), Nocardia spp. (including but not limited to Nocardia asteroides, Nocardia cyriacigeorgica and Nocardia brasiliensis), Neisseria spp. (including but not limited to Neisseria gonorrhoeae and Neisseria meningitidis), Pasteurella multocida, Plesiomonas shigelloides, Prevotella spp., Porphyromonas spp., Prevotella melaninogenica, Proteus spp.
(including but not limited to Proteus vulgaris and Proteus mirabilis), Providencia spp.
(including but not limited to Providencia alcalifaciens, Providencia rettgeri and Providencia stuartii), Pseudomonas aeruginosa, Propionibacterium acnes, Rhodococcus equi, Rickettsia spp.
(including but not limited to Rickettsia rickettsii, Rickettsia akari and Rickettsia prowazekii, Orientia tsutsugamushi (formerly: Rickettsia tsutsugamushi) and Rickettsia typhi), Rhodococcus spp., Stenotrophomonas maltophilia, Salmonella spp.
(including but not limited to Salmonella enterica, Salmonella typhi, Salmonella paratyphi, Salmonella enteritidis, Salmonella cholerasuis and Salmonella typhimurium), Serratia spp.
(including but not limited to Serratia marcesans and Serratia liquifaciens), Shigella spp. (including but not limited to Shigella dysenteriae, Shigella flexneri, Shigella boydii and Shigella sonnei), Staphylococcus spp. (including but not limited to Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus hemolyticus, Staphylococcus saprophyticus), Streptococcus spp. (including but not limited to Streptococcus pneumoniae (for example chloramphenicol-resistant serotype 4 Streptococcus pneumoniae, spectinomycin-resistant serotype 6B Streptococcus pneumoniae, streptomycin-resistant serotype 9V
Streptococcus pneumoniae, erythromycin-resistant serotype 14 Streptococcus pneumoniae, optochin-resistant serotype 14 Streptococcus pneumoniae, rifampicin-resistant serotype Streptococcus pneumoniae, tetracycline-resistant serotype 19F Streptococcus pneumoniae, penicillin-resistant serotype 19F Streptococcus pneumoniae, and trimethoprim-resistant serotype 23F Streptococcus pneumoniae, chloramphenicol-resistant serotype 4 Streptococcus pneumoniae, spectinomycin-resistant serotype 6B Streptococcus pneumoniae, streptomycin-resistant serotype 9V Streptococcus pneumoniae, optochin-resistant serotype 14 Streptococcus pneumoniae, rifampicin-resistant serotype Streptococcus pneumoniae, penicillin-resistant serotype 19F Streptococcus pneumoniae, or trimethoprim-resistant serotype 23F Streptococcus pneumoniae), Streptococcus agalactiae, Streptococcus mutans, Streptococcus pyogenes, Group A Streptococci, Streptococcus pyo genes, Group B Streptococci, Streptococcus agalactiae, Group C
Streptococci, Streptococcus anginosus, Streptococcus equismilis, Group D Streptococci, Streptococcus bovis, Group F Streptococci, Streptococcus anginosus, and Group G
Streptococci), Spirillum minus, Streptobacillus moniliformi, Treponema spp. (including but not limited to Treponema carateum, Treponema petenue, Treponema pallidum and Treponema endemicum, Tropheryma whippelii, Ureaplasma urealyticum, Veillonella spp., Vibrio spp.
(including but not limited to Vibrio cholerae, Vibrio parahemolyticus, Vibrio vulnificus, Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio alginolyticus, Vibrio mimicus, Vibrio hollisae, Vibrio fluvialis, Vibrio metchnikovii, Vibrio damsela and Vibrio fumisii), Yersinia spp. (including but not limited to Yersinia enterocolitica, Yersinia pestis, and Yersinia pseudotuberculosis) and Xanthomonas maltophilia among others.

[0075] Furthermore, other microorganisms or infectious agents that can be assayed using the methods and systems disclosed herein can comprise fungi or mold including, but not limited to, Candida spp. (including but not limited to Candida albi cans, Candida glabrata, Candida tropicalis, Candida parapsilosis, and Candida krusei), Aspergillus spp.
(including but not limited to Aspergillus fumigatous, Aspergillus flavus, Aspergillus clavatus), Cryptococcous spp. (including but not limited to Cryptococcus neoformans, Cryptococcus gattii, Cryptococcus laurentii, and Cryptococcus albidus), Fusarium spp.
(including but not limited to Fusarium oxysporum, Fusarium solani, Fusarium verticillioides, and Fusarium proliferatum), Rhizopus oryzae, Penicillium mameffei, Coccidiodes immitis, and Blastomyces dermatitidis.

[0076] Fig. 1A illustrates a front view of one embodiment of a sensor apparatus 100 for measuring a solution characteristic of a sample. The sensor apparatus 100 can comprise a sample container 104 comprising a sample chamber 108, a reference sensor 122 fabricated as part of a container cap 116 (see, e.g., Fig. 1D), and an active sensor 106 coupled to at least part of the sample chamber 108. The container cap 116 can be removably or detachably coupled or fastened to the sample container 104 (e.g., screwed or pressed on to a top of the sample container 104).

[0077] The sample container 104 can be made in part of an inert or non-conductive material. In some embodiments, the sample container 104 can comprise or be made in part of a polymeric material, a ceramic material or glass, or a combination thereof. As a more specific example, the sample container 104 can comprise or be made in part of polyvinyl chloride (PVC), poly(methyl methacrylate) (PMMA), polydimethylsiloxane (PDMS), or a combination thereof

[0078] Fig. 1B illustrates a cross-sectional side view of part of the sensor apparatus 100. Fig. 1B illustrates that the sample chamber 108 can comprise a chamber lateral wall 112 surrounding a chamber cavity 109 configured to receive a sample. The active sensor 106 can be affixed, adhered, or otherwise coupled to the chamber lateral wall 112 of the sample container 104. In other embodiments not shown in the figures, the active sensor 106 can be coupled to or otherwise positioned along a bottom of the sample container 104.

[0079] The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 at a window opening 114 defined along the chamber lateral wall 112. The chamber lateral wall 112 can comprise a recessed portion 134 surrounding the window opening 114.
The recessed portion 134 can be defined along an exterior side of the chamber lateral wall 112.

[0080] Regarding placement of the active sensor 106, the active sensor 106 can be configured such that no part of the active sensor 106 extends into the chamber cavity 109, as seen in Fig. 1C.

[0081] As will be discussed in more detail in the following sections, the active sensor 106 can be made of a conductive substrate covered in part by an active electrode layer 132.
The active electrode layer 132 of the active sensor 106 can face the chamber cavity 109 to allow the sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114.

[0082] Fig. 1C illustrates a perspective close-up view of the active sensor 106 adhered to the chamber lateral wall 112. In the embodiment shown in Fig. 1C, the active sensor 106 is adhered to the recessed portion 134 of the chamber lateral wall 112. At least part of an active electrode layer 132 of the active sensor 106 can cover a window opening defined along the chamber lateral wall 112 such that this part of the active electrode layer 132 covering the window opening 114 is positioned to be in fluid communication with the chamber cavity 109 of the sample chamber 108. When the sample chamber 108 is filled with a sample, the sample can make fluid contact with the portion of the active electrode layer 132 covering the window opening 114.

[0083] Fig. 1C also illustrates that the active sensor 106 can have its lateral sides covered by an adhesive 138. Since the active sensor 106 can comprise multiple layers, the adhesive 138 can protect certain layers of the active sensor 106 from undesired contact with the fluid sample. The adhesive 138 can act as a barrier to prevent the fluid sample from contacting the lateral sides 136 of the active sensor 106. In other embodiments not shown in the figures but contemplated by this disclosure, the recessed portion 134 of the chamber lateral wall 112 can be sized such that the active sensor 106 fits tightly within the recessed portion 134 and the walls of the recessed portion 134 adjoin or bound the lateral sides 136 of the active sensor 106. This can ensure that only the exposed portion of the active electrode layer 132 contacts the fluid sample, resulting in more accurate measurements of the solution characteristics of the fluid sample.

[0084] To adhere the active sensor 106 to the sample chamber 108, a bead of adhesive 138 can be applied to an inner ledge 140 and/or a side border 142 of the recessed portion 134 and the active sensor 106 can then be pressed into the recessed portion 134 with an end-effector of a pick-and-place machine. The active sensor 106 can be pressed or otherwise urged into the recessed portion 134 until an exterior-facing surface of the active sensor 106 is flush with an exterior surface of the chamber lateral wall 112.

[0085] The adhesive 138 can then be cured to secure the active sensor 106 in place. In some embodiments, the adhesive 138 can be a medical-grade UV-cured adhesive.
For example, the adhesive 138 can be the Dymax 1405M-T-UR-SC adhesive (curable using LED light at a wavelength of approximately 405 nm). In other embodiments, the adhesive 138 can be any low-outgassing medical grade adhesive.

[0086] As previously discussed, the active sensor 106 can be made of a conductive substrate covered in part by an active electrode layer 132. The active sensor 106 can be positioned such that the active electrode layer 132 faces the chamber cavity 109 to allow the sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114. In this embodiment, the active sensor 106 (including the active electrode layer 132) is positioned radially outward from an interior-facing or cavity-facing side of the chamber lateral wall 112 and the lateral sides 136 of the active sensor 106 are not exposed to the fluid sample.

[0087] In some embodiments, the solution characteristic measured or monitored can be a pH of the sample. When the solution characteristic measured or monitored is pH, the active electrode layer 132 can be a pH-sensitive material. For example, the pH-sensitive material can be or comprise any of silicon dioxide (SiO2), aluminum oxide (A1203), titanium dioxide (TiO2), tantalum oxide/pentoxide (Ta205), hafnium dioxide (Hf02), iridium dioxide (h02), ruthenium dioxide (RuO2), zirconium dioxide (ZrO2), or a combination thereof.

[0088] In these and other embodiments, the solution characteristic measured or monitored can be an oxidation reduction potential (ORP) of the sample. When the solution characteristic measured or monitored is the ORP of the sample, the active electrode layer 132 can be a redox-sensitive material. For example, the redox-sensitive material can be or comprise any of platinum (Pt), gold (Au), a redox sensitive metal oxide, or a combination thereof. More specifically, the redox-sensitive material can be or comprise any of silicon dioxide (SiO2), aluminum oxide (A1203), titanium dioxide (TiO2), tantalum pentoxide (Ta205), hafnium dioxide (Hf02), iridium dioxide (h02), ruthenium dioxide (RuO2), zirconium dioxide (ZrO2), or a combination thereof. Fabrication of the active sensor 106 will be discussed in more detail in later sections.

[0089] Although not shown in the figures, it is contemplated by this disclosure that the sensor apparatus 100 can be designed such that both the pH and the ORP of a sample are measured simultaneously. For example, the sample chamber 108 of the sensor apparatus 100 can comprise multiple window openings 114 defined along the chamber lateral walls 112 of the sample chamber 108. Each of these window openings 114 can then be covered by a different active sensor 106 (for example, one window opening 114 can be covered by an active sensor 106 having an active electrode layer 132 made of a redox-sensitive material and another window opening 114 can be covered by an active sensor 106 having an active electrode layer 132 made of a pH-sensitive material).

[0090] The sensor apparatus 100 can have an apparatus height. In some embodiments, the apparatus height can be between about 20.0 mm to about 50.0 mm. In other embodiments, the apparatus height can be between about 25.0 mm to about 35.0 mm. For example, the apparatus height can be about 31.3 mm.

[0091] Fig. 1D illustrates that the reference sensor 122 can be fabricated as part of a container cap 116. The reference sensor 122 can comprise a reference conduit comprising a reference conduit cavity 120 (see, e.g., Fig. 1B). The reference conduit cavity 120 can have first and second openings at opposite ends of the reference conduit cavity 120. The reference conduit 118 can be an elongate channel or passageway configured to extend into the chamber cavity 109 of the sample chamber 108.

[0092] The reference sensor 122 can also comprise a reference electrode material 149 and a wicking component 150 in fluid communication with the chamber cavity 109. The reference conduit cavity 120 can house the wicking component 150. At least some of the sample can be drawn up by the wicking component 150 in a direction of the reference electrode material 149.

[0093] The reference conduit 118 can be tapered such that a volume of the reference conduit cavity 120 tapers or narrows from a reference conduit proximal end 126 to a reference conduit distal end 128 (see, e.g., Fig. 1B). The shape of the wicking component 150 can match or accommodate the shape of the reference conduit cavity 120.
The wicking component 150 can be configured such that the shape of the wicking component 150 tapers or narrows from a wick proximal end 152 to a wick distal end 154.

[0094] The wicking component 150 can extend through a length of the reference conduit cavity 120. In some embodiments, the wicking component 150 can fill up or occupy all of the space within the reference conduit cavity 120. In other embodiments, the wicking component 150 can partially fill up or partially occupy the space within the reference conduit cavity 120.

[0095] At least part of the wicking component 150 can be in fluid communication with the chamber cavity 109 of the sample chamber 108 such that when the sample chamber 108 is filled with the sample, at least some of the sample in the sample chamber 108 is drawn up, absorbed, or otherwise wicked by at least a portion of the wick distal end 154 in a direction of the wick proximal end 152. The wicking component 150 can be made of a polymeric material that draws up the fluid sample towards the reference electrode material 149 by capillary action.

[0096] In some embodiments, at least part of the wick distal end 154 can extend past the reference conduit second opening such that the wick distal end 154 protrudes or extends into the chamber cavity 109 of the sample chamber 108. In these embodiments, the wick distal end 154 can extend or protrude into the sample when the sample chamber 108 is filled by the sample.

[0097] In other embodiments, the wick distal end 154 is positioned proximal or above the reference conduit second opening such that the wick distal end 154 does not protrude or extend into the chamber cavity 109 of the sample chamber 108. In these embodiments, the wick distal end 154 can still be in fluid communication with the sample chamber 108 and the fluid sample can still reach or contact the wick distal end 154 by being drawn up into the reference conduit 118 by capillary action or by perturbing or shaking the sample container 104.

[0098] As previously discussed, the wicking component 150 can be made in part of a porous material. The wicking component 150 can be made in part of a material comprising pores sized between 15 um to about 150 um (e.g., about 50 um). In some embodiments, the wicking component 150 can be made in part of a polymeric material. As a more specific example, the wicking component 150 can be made in part of a porous polymeric material comprising pores sized between 15 um to about 150 um. In one embodiment, the wicking component 150 can be made in part of high-density polyethylene (HDPE).
For example, the wicking component 150 can be made in part of HDPE having pores sized about 50 um. In other embodiments, the wicking component 150 can be made in part of natural fibers. For example, the wicking component 150 can be made in part of cellulose fibers, pulp, paper, cotton, or a combination thereof.

[0099] The wicking component 150 can also be treated by a surfactant such that at least a surface of the wicking component 150 is covered by the surfactant. In some embodiments, the wicking component 150 can be saturated by the surfactant or immersed in a solution comprising the surfactant prior to being introduced into the reference conduit cavity 120. The surfactant can be configured to increase a hydrophilicity of the wicking component 150 (i.e., to make a substantially hydrophobic surface of the wicking component 150 more hydrophilic). In some embodiments, the surfactant can be a fluorosurfactant. In other embodiments, the surfactant can be a non-ionic surfactant such as one or more Poloxamers. As a more specific example, the surfactant can comprise Pluronic0 F-68.

[0100] In one embodiment, the reference conduit 118 can be substantially shaped as a conic or frustoconic having a reference conduit cavity 120 also substantially shaped as a conic or frustoconic. In other embodiments, the reference conduit 118 can be substantially shaped as an elongate pyramid having a polygonal-shaped base. For example, the reference conduit 118 can be substantially shaped as an elongate triangular pyramid, square pyramid, or a pentagonal pyramid. In additional embodiments, the reference conduit 118 can be substantially shaped as a cylinder having a substantially cylindrical-shaped reference conduit cavity 120. In these embodiments, the reference conduit 118 can have a tapered reference conduit distal end 128 (see, e.g., Fig. 1B).

[0101] As shown in Fig. 1D, at least part of the wicking component 150 can be in fluid contact with the sample in the sample chamber 108. At least some of the sample can be drawn up by the wicking component 150 in a direction of the wick proximal end 152. The reference electrode material 149 can be disposed at the wick proximal end 152.
Fig. 1D
also illustrates that at least part of the active electrode layer 132 can be in fluid contact with the sample in the sample chamber 108. When the wicking component 150 draws or wicks up the sample, the sample can reach the reference electrode material 149 and charge carriers within the sample can establish an electrical connection between the reference electrode material 149 of the reference sensor 122 and the active electrode layer 132 of the active sensor 106. When both the reference sensor 122 and the active sensor 106 are electrically coupled to a reader apparatus 190, the reader apparatus 190 can be used to measure a solution characteristic of the sample.

[0102] The solution characteristic of the sample can be determined based on a potential difference measured between the active sensor 106 and the reference sensor 122 when the reference sensor 122 and the active sensor 106 are electrically coupled to the reader apparatus 190. For example, the reference sensor 122 can provide a stable half-cell potential compared to the active sensor 106 when both the reference electrode material 149 and the active electrode layer 132 are in fluid contact with the sample within the sample chamber 108.

[0103] In some embodiments, the reference electrode material 149 can be an electrically-conductive ink applied or dispensed on the wick proximal end 152.
The electrically-conductive ink applied or dispensed on the wick proximal end 152 can be hardened by curing. More specifically, the electrically-conductive ink can be a silver-silver chloride (Ag-AgC1) ink.

[0104] At least part of the reference electrode material 149 can be coupled to the wicking component 150. For example, the reference electrode material 149 can be a cured and hardened mass positioned at the wick proximal end 152. In certain embodiments, the reference electrode material 149 can be positioned in the middle of the container cap 116.
In some embodiments, at least part of the reference electrode material 149 can protrude or extend beyond the container cap 116.

[0105] One advantage of the wicking component 150 disclosed herein is that the wicking component 150 can draw up the sample and the sample can advance by capillary action through the pores of the wicking component 150 toward the reference electrode material 149. For example, the liquid sample can be wicked to the wick proximal end 152 where it makes fluid contact with the reference electrode material 149. When the reference electrode material 149 is made of a material such as silver-silver chloride (Ag-AgC1), the wicking component 150 can act as a barrier or hindrance to silver ions (AO
that would otherwise diffuse freely into the sample within the sample chamber 108. Such silver ions can be harmful to or otherwise affect the growth of the microorganisms or infectious agents in the sample. The wicking component 150 can act as a barrier or hindrance to the harmful silver ions by slowing down or stalling the diffusion of such ions into the sample. The wicking component 150 having the dimensions and shape disclosed herein can be effective in slowing down or stalling the diffusion of such harmful ions.

[0106] When the reference sensor 122 is implemented as a container cap 116, the container cap 116 can have dimensions as defined by a cap width (or diameter) and a cap height. In some embodiments, the cap width can be between about 10.0 mm to about 20.0 mm. For example, the cap width can be about 15.7 mm. In some embodiments, the cap height can be between about 5.0 mm to about 20.0 mm. For example, the cap height can be about 10.5 mm. When the container cap 116 is fastened, affixed, or otherwise coupled to the sample container 104, the sensor apparatus 100 can have an apparatus height as measured from a bottom of the sample container 104 to a cap top 130 of the container cap 116.

[0107] The wicking component 150 can have a wick height as measured from the wick proximal end 142 to the wick distal end. In some embodiments, the wick height can be between about 10.0 mm to about 20.0 mm. More specifically, the wick height can be between about 14.0 mm to about 15.0 mm. For example, the wick height can be about 14.8 nam.

[0108] As illustrated in Fig. 1D, the reference electrode material 149 can be positioned or disposed, at least partially, within a divot, depression, or concave region in a center of the container cap 116 above the wicking component 150. When the reference sensor 122 is a cured or hardened electrically-conductive ink or solution (e.g., Ag-AgC1 ink), the divot, depression, or concave region can act as a receiving space for the liquid ink or solution to be cured.

[0109] In some embodiments, the reference electrode material 149 can have a reference electrode height and a reference electrode width. The reference electrode height can be between about 0.2 mm and 1.0 mm. For example, the reference electrode height can be about 0.4 mm. The reference electrode width can be between about 2.0 mm to about 5.0 mm. For example, the reference electrode width can be about 3.0 mm. One advantage of the reference sensor 122 disclosed herein is that the reference sensor 122 can act as a stable reference electrode or provide a stable reference potential for up to 10-hours of testing or operation.

[0110] Fig. 1D also illustrates that the sensor apparatus 100 can comprise an aeration port 160 defined along a bottom side of the sample chamber 108. In other embodiments not shown in the figures, the aeration port 160 can be defined along the chamber lateral wall 112 of the sample chamber 108.

[0111] The aeration port 160 can be covered by a first air-permeable membrane. The aeration port 160 and the first air-permeable membrane can be configured to allow a gas 162 to enter the sample chamber 108.

[0112] In some embodiments, the gas 162 can be ambient air (e.g., the air in a laboratory, clinical setting, or testing facility). In other embodiments, the gas 162 can comprise a combination of pressurized oxygen, carbon dioxide, nitrogen, and argon.
Aerating the sample can accelerate the growth of a microbial population within the sample by providing an oxygen rich environment within the sample chamber 108.

[0113] In alternative embodiments not shown in the figures, the aeration port 160 can be defined along a cap top 130 of the container cap 116 and the gas 162 can be pumped into the sample chamber 108 from the top of the sample container 104.

[0114] The gas 162 (e.g., ambient air) can be pumped into the sample chamber 108 by a micropump or another pump-type device integrated within the reader apparatus 190. The gas 162 (e.g., ambient air) can be pumped or otherwise directed into the sample chamber 108 through the aeration port 160 and the first air-permeable membrane at a constant flow rate of between about 1.0 and 10.0 mL/min. In other embodiments, the gas 162 (e.g., ambient air) can be pumped or otherwise directed into the sample chamber 108 through the aeration port 160 and the first air-permeable membrane at specific duty cycles or intervals.

[0115] In certain embodiments, a second air-permeable membrane can cover at least part of an underside of the container cap 116. The second air-permeable membrane can allow any gas 162 pumped or otherwise introduced into the sample chamber 108 to exit the sample chamber 108 while also preventing any liquid within the sample chamber 108 from spilling out of the sample container 104.

[0116] In some embodiments, the first air-permeable membrane and the second air-permeable membrane can be made of the same material. The first air-permeable membrane and the second air-permeable membrane can be made of a hydrophobic air-permeable film or thin-sheet. For example, the first air-permeable membrane and the second air-permeable membrane can both be made of or comprise polytetrafluoroethylene (PTFE).

[0117] As shown in Fig. 1D, the container cap 116 can be removably or detachably coupled or fastened to the sample container 104 by being screwed on to a proximal portion of the sample container 104 via a threaded connection 164. When the container cap 116 (serving as part of the reference sensor 122) is fastened or coupled to the sample container 104 by the threaded connection 164, an airflow pathway 166 can be created as air enters the aeration port 160 through the first air-permeable membrane into the sample chamber 108. The air then exits the sample chamber 108 through the second air-permeable membrane and air gaps 168 defined in between the threads of the container cap 116 and the sample container 104.

[0118] The container cap 116 can be made in part of a transparent or clear material or a transparent or clear non-conducting material. In other embodiments, the container cap 116 can be made in part of a translucent or see-through material. For example, at least part of the wicking component 150 can be visible through the sides of the container cap 116. This can allow a user or operator of the sensor apparatus 100 to observe the wicking of the fluid sample from the wick distal end 154 to the wick proximal end 152 when the container cap 116 is fastened to the sample container 104 and ensure that at least some of the sample is able to reach the reference electrode material 149 at the wick proximal end 152. In some embodiments, the container cap 116 can be made in part of a clear or transparent polymeric material, glass, or a combination thereof.

[0119] In some embodiments, the sample container 104, the container cap 116, or a combination thereof can be made in part of an inert polymeric material. For example, the sample container 104, the container cap 116, or a combination thereof can be made in part of at least one of polyoxymethylene, polyamide, polyethylene, acrylonitrile butadiene styrene, polycarbonate, polypropylene, or co-polymers or composites thereof.
In other embodiments, the sample container 104, the container cap 116, or a combination thereof can be made in part a glass material such as borosilicate glass or a ceramic material.

[0120] Fig. 2 illustrates that the active sensor 106 can also be insert molded into part of the chamber lateral wall 112 when the sample container 104 is made of a polymeric material. For example, the active sensor 106 can be insert-molded into the chamber lateral wall 112 while the sample container 104 is being formed by injection molding.

[0121] When the active sensor 106 is inserted molded into part of the chamber lateral wall 112 of the sample chamber 108, the active sensor 106 can have its lateral sides 136 encapsulated by the polymeric material used to make the chamber lateral wall 112.

[0122] In the embodiment shown in Fig. 2, the active sensor 106 can be insert molded such that the active electrode layer 132 faces the chamber cavity 109 to allow the sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114.

[0123] Figs. 3A and 3B are black-and-white images of an active sensor 106 insert molded into a polymeric material representing the material used to construct the chamber lateral wall 112 of the sample chamber 108 (see, e.g., Figs. 1A-1D). In some embodiments, the sample chamber 108 can be made in part of an inert polymeric material such as polyoxymethylene, polyamide, polyethylene, acrylonitrile butadiene styrene, polycarbonate, or polypropylene.

[0124] Fig. 3A shows a top plan view of the side of the active sensor 106 covered by the active electrode layer 132. As previously discussed, the active sensor 106 can be insert molded such that the active electrode layer 132 faces the chamber cavity 109 to allow the sample within the chamber cavity 109 (see, e.g., Fig. 1D) to be in fluid contact with an exposed region of the active electrode layer 132.

[0125] Fig. 3B shows a top plan view of a side of the active sensor 106 opposite the active electrode layer 132. The side of the active sensor 106 shown in Fig. 3B
can be used to contact the conductive connections of the reader apparatus 190 (see, e.g., Figs. 14 and 15). As will be discussed in more detail in the following sections, this side of the active sensor 106 can be referred to as a conductive layer.

[0126] As shown in Figs. 3A and 3B, the lateral sides 136 of the active sensor 106 can be encapsulated by the polymeric material. This can ensure that only the exposed portion of the active electrode layer 132 contacts the fluid sample within the sample chamber 108, resulting in more accurate measurements of the solution characteristics of the fluid sample.

[0127] Although not shown in the figures, it is contemplated by this disclosure that the active sensor 106 can be affixed or otherwise coupled to the chamber lateral wall 112 by focally melting (e.g., by ultrasonic welding) a portion of the chamber lateral wall 112 surrounding the window opening 114 (see, e.g., Figs. 1B-1D for the location of the window opening 114) and pressing the active sensor 106 onto the melted portion of the chamber lateral wall 112. Once the melted portion of the chamber lateral wall 112 cools, the active sensor 106 is now affixed or coupled to the chamber lateral wall 112.

[0128] Fig. 4A illustrates a perspective view of one embodiment of an active sensor 106 with two of its lateral sides 136 visible. As shown in Fig. 4A, the active sensor 106 can be substantially shaped as a flattened or truncated rectangular prism. In other embodiments, the active sensor 106 can be substantially disk-shaped or shaped as a flattened or truncated polygonal prism (e.g., a flattened or truncated pentagonal prism or hexagonal prism).

[0129] Fig. 4A also illustrates that when the active sensor 106 is substantially shaped as a rectangular prism, the active sensor 106 can have a sensor length dimension 400, a sensor width dimension 402, and a sensor height dimension 404. In some embodiments, the sensor length dimension 400 can be between about 100 um and 6.0 mm, the sensor width dimension 402 can be between about 100 um and 6.0 mm, and the sensor height dimension 404 can be between about 10 um and 0.70 mm. For example, when the active sensor 106 is substantially shaped as a rectangular prism, the active sensor 106 can have a sensor length dimension 400 of about 6.0 mm, a sensor width dimension 402 of about 6.0 mm, and a sensor height dimension 404 of about 0.61 mm.

[0130] Fig. 4B illustrates a side view of one embodiment of an active sensor 106 used for measuring an oxidation reduction potential (ORP) of a sample. In this embodiment, the active sensor 106 can have an active electrode layer 132 made of a noble metal. For example, the active electrode layer 132 can be made of platinum, gold, or a combination or composite thereof.

[0131] The active electrode layer 132 can be adhered to one side of a conductive substrate 406 via an adhesion layer 408. The conductive substrate 406 can be made of a conductive material such as stainless steel (SS). For example, the conductive substrate 406 can be SS 316. In other embodiments, the conductive substrate 406 can be made of aluminum, copper, or any combination or composite of aluminum, copper, or stainless steel.

[0132] In some embodiments, the adhesion layer 408 can be a thin layer of chromium (Cr). Alternatively, the adhesion layer 408 can be a thin layer of gold, nickel, titanium or tantalum. The adhesion layer 408 can be disposed in between the conductive substrate 406 and the active electrode layer 132.

[0133] In alternative embodiments, the active electrode layer 132 can be deposited directly onto one side of the conductive substrate 406 without an adhesion layer 408.

[0134] The active electrode layer 132 can have an active electrode layer thickness 410 of between about 50 nm and 500 nm (e.g., about 400 nm). The adhesion layer 408 can have an adhesion layer thickness 412 of between about 5 nm and 50 nm (e.g., about 20 nm). A
ratio of the adhesion layer thickness 412 to the active electrode layer thickness 410 can be between about 1:10 and 1:20.

[0135] The conductive substrate 406 can have a substrate layer thickness 414. The substrate layer thickness can be between about 10 um and 0.70 mm (e.g., about 0.61 mm).

[0136] Fig. 4C illustrates a side view of another embodiment of an active sensor 106 used for measuring a pH of a sample. In this embodiment, the active sensor 106 can have an active electrode layer 132 made of a pH-sensitive material. The pH-sensitive material can be deposited as a layer directly onto the conductive substrate 406 or via an adhesion layer 408.

[0137] For example, the active electrode layer 132 can be made of a metal oxide. For example, the active electrode layer 132 can be made of tantalum pentoxide (Ta205). In other embodiments, the active electrode layer 132 can be made of silicon dioxide (SiO2), silicon nitride (Si3N4), aluminum oxide (A1203), titanium dioxide (TiO2), hafnium dioxide (Hf02), iridium dioxide (Ir02), ruthenium dioxide (RuO2), zirconium dioxide (ZrO2), or a combination or composite thereof.

[0138] The conductive substrate 406 can be made of a conductive material such as stainless steel (SS). For example, the conductive material can be SS 316. In other embodiments, the conductive substrate 406 can be made of aluminum, copper, or any combination or composite of aluminum, copper, or stainless steel.

[0139] In some embodiments, the adhesion layer 408 can be a thin layer of chromium (Cr). Alternatively, the adhesion layer 408 can be a thin layer of gold, nickel, titanium or tantalum. The adhesion layer 408 can be disposed in between the conductive substrate 406 and the active electrode layer 132.

[0140] As previously discussed, the active electrode layer 132 can be deposited directly onto the conductive substrate 406 without an adhesion layer 408.

[0141] The active electrode layer 132 can have an active electrode layer thickness 410 of between about 50 nm and 500 nm (e.g., about 400 nm). The adhesion layer 408 can have an adhesion layer thickness 412 of between about 5 nm and 50 nm (e.g., about 20 nm). A
ratio of the adhesion layer thickness 412 to the active electrode layer thickness 410 can be between about 1:10 and 1:20.

[0142] The conductive substrate 406 can have a substrate layer thickness 414. The substrate layer thickness can be between about 10 um and 0.70 mm (e.g., about 0.61 mm).

[0143] Fig. 4D illustrates a side view of another embodiment of an active sensor 106 used for measuring a pH of a sample. In this embodiment, a surface modification technique can be used to modify a platinum layer 416. For example, an oxygen plasma treatment can be used to oxidize the platinum layer 416 to create a platinum oxide/dioxide (Pt02) layer 418. The platinum oxide layer 418 thus formed can respond to hydrogen ions and be used as a pH-sensitive layer. In this manner, the platinum oxide layer 418 can act as the active electrode layer 132.

[0144] The platinum layer 416 can be adhered to a conductive substrate 406 via an adhesion layer 408. The conductive substrate 406 can be made of a conductive material such as stainless steel (SS). For example, the conductive substrate can be SS
316. In other embodiments, the conductive substrate 406 can be made of aluminum, copper, or any combination or composite of aluminum, copper, or stainless steel.

[0145] In some embodiments, the adhesion layer 408 can be a thin layer of chromium (Cr). Alternatively, the adhesion layer 408 can be a thin layer of gold, nickel, titanium or tantalum. The adhesion layer 408 can be disposed in between the conductive substrate 406 and the active electrode layer 132.

[0146] In alternative embodiments, the platinum layer 416 can be deposited directly onto one side of the conductive substrate 406 without an adhesion layer 408.

[0147] The platinum layer 416 can have a layer thickness of between about 50 nm and 500 nm (e.g., about 400 nm). The adhesion layer 408 can have an adhesion layer thickness 412 of between about 5 nm and 50 nm (e.g., about 20 nm).

[0148] The conductive substrate 406 can have a substrate layer thickness 414. The substrate layer thickness can be between about 10 um and 0.70 mm (e.g., about 0.61 mm).

[0149] The platinum oxide layer 418 can have an oxide layer thickness 420.
The oxide layer thickness 420 can be between about 10 nm and 100 nm.

[0150] As previously discussed, the deposited layers can be selected to achieve a certain desired sensitivity or specificity towards a particular analyte. Other surface modification techniques such as self-assembled monolayers (SAMs), bio-functionalization with antibodies, binding antibody fragments, binding aptamers, binding DNA, and plasma treatments can also be employed to alter the surface properties of the deposited layers and thereby tune their specificity and sensitivity.

[0151] Fig. 5A illustrates a side view of yet another embodiment of an active sensor 106. This embodiment of the active sensor 106 leverages the scale and efficiency of printed circuit board (PCB) manufacturing techniques.

[0152] The active sensor 106 can be made of a non-conductive PCB substrate covered in part by an active electrode layer 132. In some embodiments, the non-conductive PCB substrate 500 can be made of polyimide. In other embodiments, the non-conductive PCB substrate 500 can be made of a glass-reinforced epoxy laminate material such as an FR-4 composite material. In certain embodiments, the PCB substrate 500 can be a flexible PCB material.

[0153] In some embodiments, the active electrode layer 132 can be made of a noble metal. For example, the active electrode layer 132 can be made of platinum (see, e.g., Figs.
5A, 5B, and 6A-6C), gold (see, e.g., Figs. 6A-6C), or a combination or composite thereof.
The platinum or gold can be electrodeposited or sputter deposited on the PCB
substrate 500.

[0154] The active electrode layer 132 can have an active electrode layer thickness of at least 50 nm. In certain embodiments, the active electrode layer 132 can have an active electrode layer thickness of at least 400 nm. When the active electrode layer 132 is made of platinum, the active sensor 106 can be used for measuring or monitoring the ORP of a sample.

[0155] In an alternative embodiment, a platinum layer deposited on the non-conductive PCB substrate 500 can be modified with a surface modification technique to turn the platinum layer into a pH-sensitive layer (see, e.g., Fig. 4D). For example, an oxygen plasma treatment can be used to oxidize the platinum layer to create a platinum oxide (Pt02) layer. The platinum oxide layer thus formed can respond to hydrogen ions and be used as a pH-sensitive layer. In this embodiment, the active sensor 106 can be used to measure or monitor the pH of a sample.

[0156] The PCB substrate 500 can be patterned with conductive contacts or a conductive contact layer 502 on a side of the substrate opposite the active electrode layer 132. In some embodiments, the conductive contact layer 502 can be a gold layer. In other embodiments, the conductive contact layer 502 can be made of another type of conductive metal such as platinum, nickel, copper, or alloys or composites thereof.

[0157] As shown in Fig. 5A, the active electrode layer 132 can be electrically coupled to the conductive contacts or conductive contact layer 502 by one or more conductive vias 504. In one embodiment, the conductive vias 504 can be made in part of copper or a copper alloy. In other embodiments, the conductive vias 504 can be made of another type of conductive metal such as gold.

[0158] In some embodiments, each active sensor 106 can have at least one conductive via 504 positioned in a center of the sensor package. In other embodiments, the conductive via 504 can be positioned near a periphery or edge of the sensor package.

[0159] The conductive vias 504 can be formed by electroplating, deposition, or a combination thereof. Moreover, additional features or patterns can be formed on the PCB
substrate 500 using standard PCB etching processes.

[0160] Fig. 5B illustrates a single PCB board covered by an active electrode layer 132 (e.g., platinum) that can be singulated into numerous individual active sensors 106. For example, one PCB board can be singulated to produce between 400 and 500 active sensors 106.

[0161] Fig. 6A is a black-and-white image showing an active sensor 106 comprising three individual active electrodes including a gold (Au) active electrode 600A, a first platinum (Pt) active electrode 600B, and a second Pt active electrode 600C.
The active sensor 106 can be produced using the PCB manufacturing techniques previously discussed.
The only difference being that different active electrode materials (e.g., Au and Pt) were electroplated or deposited on the same non-conductive PCB substrate 500. For example, one section or strip of the non-conductive PCB substrate 500 can be covered by a first active electrode material or layer (e.g., Au) and other sections or strips of the same non-conductive PCB substrate 500 can be covered by a second active electrode material or layer (e.g., Pt).

[0162] As shown in Fig. 6A, the active sensor 106 can be coupled (e.g., adhered or insert molded) to a part of the lateral chamber wall 112 of the sample chamber 108. Since the sample container 104 shown in Fig. 6A is fabricated as a clear container, the active electrode side of the active sensor 106 is visible through the clear walls of the sample chamber 108. When the sample chamber 108 is filled with a fluid sample (not shown in Fig. 6A), the fluid sample can contact the gold active electrode 600A, the first platinum active electrode 600B, and the second platinum active electrode 600C through the window opening 114 defined along the chamber lateral wall 112.

[0163] One advantage of an active sensor 106 comprising multiple active electrodes is that each electrode can report a unique potential with respect to the same reference electrode or reference sensor (e.g., the reference sensor 122). Moreover, the active electrodes can be made of different materials such that different solution characteristics (e.g., ORP and pH) of the sample can be measured or monitored at the same time.

[0164] Although not shown in the figures, it is contemplated by this disclosure that an active sensor 106 made of numerous active electrodes arranged as an active electrode array (e.g., a 96 electrode array) can be integrated into a single sensor apparatus 100 to measure multiple solution characteristics of a sample at the same time. The multiple active electrodes can be patterned on the non-conductive PCB substrate 500 using techniques common in the PCB industry including selective etching, photoresist layers, shadow masking, or a combination thereof.

[0165] Fig. 6B is a black-and-white image showing a close-up view of a contact side of the active sensor 106 shown in Fig. 6A. For example, as shown in Fig. 6B, each of the active electrodes can have its own conductive contact strip 602 or section deposited or plated on an opposite side of the non-conductive PCB substrate 500. For example, the conductive contact strips 602 or sections can be made of gold. The conductive contact strips 602 or sections can be segmented instances of the conducive contact layer 502 (see, e.g., Fig. 5A). The active electrodes can be electrically coupled to the conductive contacts by conductive vias (not shown in Fig. 6B) extending through the non-conductive PCB
substrate 500.

[0166] Fig. 6C is a black-and-white image showing a single PCB board that can be singulated into individual active sensors 106 (see, e.g., Figs. 6A and 6B). As previously discussed, a non-conductive PCB board can be covered on one side by an active electrode layer 132 using electroplating or sputter deposition. The other side of the PCB board can be covered in part by a conductive contact layer 502 (also via electroplating or a deposition technique). The active electrode layer 132 can be electrically coupled to the conductive contact layer 502 by conductive vias 504 extending through the non-conductive PCB
board. As previously discussed, one PCB board processed in this manner can be singulated to produce between 400 and 500 active sensors 106.

[0167] Fig. 7 illustrates yet another embodiment of an active sensor 106 made by covering a non-conductive polymeric substrate 700 comprising a through-hole 702 with an active electrode layer 132 and a conductive contact layer 502.

[0168] The non-conductive polymeric substrate 700 can be a substrate made of any type of injection-molded plastic such as polyamide, polycarbonate, polyoxymethylene, polystyrene, acrylonitrile butadiene styrene, polypropylene, polyethylene, or copolymers or blends thereof.

[0169] In some embodiments, the active electrode layer 132 is a noble metal layer. For example, the active electrode layer 132 can be a layer of platinum, a layer of gold, or a combination or composite thereof. The platinum or gold layer can be deposited or otherwise applied to the conductive polymeric substrate 700 via sputter deposition (e.g., physical vapor deposition (PVD) sputter deposition), evaporation deposition, or electrodeposition. In some embodiments, the platinum or gold layer can be printed using screen printing.

[0170] The active electrode layer 132 can have an active electrode layer thickness of at least 50 nm. In certain embodiments, the active electrode layer 132 can have an active electrode layer thickness of at least 400 nm. When the active electrode layer 132 is made of platinum or gold, the active sensor 106 can be used for measuring or monitoring the ORP
of a sample.

[0171] In an alternative embodiment, a platinum layer deposited on the non-conductive polymeric substrate 700 can be modified with a surface modification technique to turn the platinum layer into a pH-sensitive layer (see, e.g., Fig. 4D). For example, an oxygen plasma treatment can be used to oxidize the platinum layer to create a platinum oxide (Pt02) layer. The platinum oxide layer thus formed can respond to hydrogen ions and be used as a pH-sensitive layer. In this embodiment, the active sensor 106 can be used to measure or monitor the pH of a sample.

[0172] In some embodiments, the conductive contact layer 502 can be a gold layer. In other embodiments, the conductive contact layer 502 can be made of another type of conductive metal such as platinum, nickel, copper, or alloys or composites thereof.

[0173] The through-hole 702 can have a diameter between about 10 um to 100 um. In some embodiments, the active sensor 106 can have a width dimension of between about 100 um and 6.0 mm and a length dimension of between about 100 um and 6.0 mm.
For example, the active sensor 106 can have a width dimension of about 100 um and a length dimension of about 100 um.

[0174] Figs. 8A and 8B are side cross-sectional views illustrating two different embodiments of the active sensor 106. In both embodiments, the ends of the through-hole 702 are covered by the active electrode layer 132 and the conductive contact layer 502. As shown in Figs. 8A and 8B, a conductive coating can cover the lateral sides of the through-hole 702.

[0175] In the embodiment shown in Fig. 8A, the conductive coating is comprised of the same material as the active electrode layer 132. In the embodiment shown in Fig. 8B, the conductive coating is comprised of the same material as the conductive contact layer 502.
Whether the lateral sides of the through-hole 702 are covered by the active electrode material or the conductive coating material can be determined by which layer is first deposited on the non-conductive polymeric substrate 700.

[0176] As a more specific example, the conductive coating covering the lateral sides of the through-hole can be a coating of platinum when the active electrode layer 132 is a layer of platinum and the layer of platinum is first deposited on the non-conductive polymeric substrate 700. Alternatively, the conductive coating covering the lateral sides of the through-hole can be a coating of gold when the conductive contact layer 502 is a layer of gold and the layer of gold is first deposited on the non-conductive polymeric substrate 700.

[0177] In some embodiments (for example, as shown in Figs. 8A and 8B), the entire through-hole 702 does not need to be filled as long as the lateral sides of the through-hole 702 are covered by the conductive coating. The conductive coating can serve as an electrical connection or conductive path between the two sides of the active sensor 106.
Alternatively, at least part of the through-hole 702 can be filled with the conductive coating.

[0178] In some embodiments, the non-conductive polymeric substrate 700 can start off as a sheet of plastic having an array of small through-holes 702 defined throughout the sheet of plastic. The sheet of plastic can then be covered first with the active electrode layer 132 or the conductive contact layer 502. The lateral sides of the through-holes 702 and at least one of the ends of the through-holes 702 can then be coated by the material used to initially cover the sheet of plastic. The other side of the sheet of plastic including the remaining open ends of the through-holes 702 can then be covered by the conductive contact layer 502 or the active electrode layer 132, depending on which layer went first.
Once the sheet of plastic is covered on both sides, the sheet of plastic can be singulated to produce the individual active sensors 106. Active sensors 106 produced using this method can be made as small as 100 um by 100 um (W x L).

[0179] Fig. 9 illustrates that a large sheet of non-conductive plastic or a large PCB can be processed using the methods disclosed herein (covered by an active electrode layer, an adhesion layer, a conductive layer, or a combination thereof) and then singulated into numerous active sensors 106. In some embodiments, the large sheet of non-conductive plastic or the large PCB can be singulated using sawing, laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or a combination thereof.

[0180] Fig. 10 illustrates a side view of another embodiment of a sensor apparatus 100 comprising an active sensor 106 made of a conductive dowel 1000. In some embodiments, the conductive dowel 1000 can be a stainless steel dowel. The conductive dowel 1000 can be covered in part by an active electrode layer 132 extending into the chamber cavity 109.
The part of the conductive dowel 1000 covered by the active electrode layer 132 can extend into the chamber cavity 109 to allow the sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132.

[0181] The conductive dowel 1000 can be coupled to at least part of the chamber lateral wall 112 at a window opening defined along the chamber lateral wall 112. An opposite end of the conductive dowel 1000 (the end not covered by the active electrode layer 132) can extend out of the chamber lateral wall 112 to contact one or more conductive connections of a reader apparatus 190. The conductive dowel 1000 can be shaped substantially as a cylinder having rounded edges.

[0182] The conductive dowel 1000 can be insert molded or adhesive bonded to the chamber lateral wall 112.

[0183] In some embodiments, the active electrode layer 132 is a noble metal layer. For example, the active electrode layer 132 can be a layer of platinum, a layer of gold, or a combination or composite thereof. The platinum or gold layer can be deposited or otherwise applied to the conductive dowel 1000 via sputter deposition (e.g., physical vapor deposition (PVD) sputter deposition), evaporation deposition, or electrodeposition.

[0184] The active electrode layer 132 can have an active electrode layer thickness of at least 50 nm. In certain embodiments, the active electrode layer 132 can have an active electrode layer thickness of at least 400 nm. When the active electrode layer 132 is made of platinum or gold, the active sensor 106 can be used for measuring or monitoring the ORP
of a sample.

[0185] In an alternative embodiment, a platinum layer deposited on the conductive dowel 1000 can be modified with a surface modification technique to turn the platinum layer into a pH-sensitive layer. For example, an oxygen plasma treatment can be used to oxidize the platinum layer to create a platinum oxide (Pt02) layer. The platinum oxide layer thus formed can respond to hydrogen ions and be used as a pH-sensitive layer. In this embodiment, the active sensor 106 can be used to measure or monitor the pH of a sample.

[0186] Fig. 11 is a graph illustrating a change in the oxidation reduction potential (ORP) of three samples containing E. coli ATCC 25922 measured over time using three different sensors. As shown in Fig. 11, one sensor is a traditional ORP probe that is commonly used in diagnostic settings (for example, the commercially-available ORP probe distributed by Mettler-Toledo, LLC). The other two sensors are embodiments of the sensor apparatus 100 disclosed herein with one having an active sensor 106 comprising a platinum active electrode layer deposited by evaporation deposition and the other having an active sensor 106 comprising an electroplated platinum active electrode layer. The active sensors 106 in both embodiments are coupled to at least part of the chamber lateral wall 112 at a window opening 114 defined along the chamber lateral wall 112. In these embodiments, the active sensors 106 are positioned such that no part of the active sensors 106 extends into the chamber cavity 109 of the sample chamber 108. The change in ORP was measured by a reader apparatus 190 when each of the two sensor apparatus 100 was placed within the reader apparatus 190.

[0187] As shown by the three E. coli growth curves, the two sensor apparatus 100 performed similar to the commercially-available ORP probe. Any variations in the signal response were within acceptable ranges.

[0188] Fig. 12 is a graph illustrating a change in the pH of four samples containing different starting concentrations of E. coli ATCC 25922 measured over time using the sensor apparatus 100 disclosed herein having an active sensor 106 comprising a tantalum oxide/pentoxide (Ta205) active electrode layer.

[0189] As shown in Fig. 12, the E. coli growth curves measured followed the classical growth pattern of bacteria having a lag phase at the outset, following by an exponential phase, and ending in a stationary phase. The pattern or shape of the curves can be attributed to cellular activity undertaken by the active E. coli within the samples.

[0190] Fig. 13A illustrates a perspective view of a reader apparatus 190 configured to determine a solution characteristic of the sample within the sample chamber 108 of the sensor apparatus 100. The reader apparatus 190 can determine the solution characteristic of the sample based on a potential difference measured between the active sensor 106 (more specifically, the active electrode layer 132) and the reference sensor 122 (more specifically, the reference electrode material 149) when the active sensor 106 and the reference sensor 122 are electrically coupled via conductive connections or interfaces within the reader apparatus 190. The reader apparatus 190 can act as a voltmeter or another type of high-impedance amplifier or sourcemeter to measure relative changes in an equilibrium potential at an interface between the electrode layers in fluid contact with a sample containing electro-active redox species or charged ions.

[0191] The solution characteristic of the sample can change as the amount of electro-active redox species or the amount of IT ions changes due to the growth or metabolism (or lack thereof) of infectious agents within the sample. For example, the amount of electro-active redox species in the sample can change as a result of cellular activity undertaken by the infectious agents. As a more specific example, the amount of oxygen and the amount of electron donors can change as the amount of energy carriers, such as nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FADH2), changes due to the growth or metabolism (or lack thereof) of infectious agents within the sample.

[0192] The reader apparatus 190 can measure the oxidation reduction potential (ORP) of a sample when the active electrode layer 132 of the sensor apparatus 100 is made of a redox-sensitive material such as platinum (Pt) or gold (Au). Moreover, the reader apparatus 190 can also measure the pH of a sample when the active electrode layer 132 of the sensor apparatus 100 is made of a pH-sensitive material such as a metal-oxide layer.

[0193] Fig. 13A illustrates that the reader apparatus 190 can comprise a reader housing 1300 configured to house certain functional components of the reader apparatus including a main controller 1301 (see, e.g., Fig. 13C), a signal readout control unit 1303 (see, Figs. 14, 15A, and 15B), a thermal control module 1305 (see, e.g., Figs.
13B, 13C, and 13D), and an aeration control module 1307 (see, e.g., Fig. 13B and 13C).
The reader housing 1300 can also expose a touchscreen display 1302 configured to display measurement results and allow a user to input commands to the reader apparatus 190.

[0194] A lid 1304 or cover of the reader apparatus 190 can be be opened or lifted up to reveal a container receiving space 1306 (see, e.g., Fig. 13B) configured to accommodate or receive the sensor apparatus 100 for analysis or investigation by the reader apparatus 190.

[0195] Fig. 13B illustrates a partial cutaway view of the reader apparatus 190 with a sample-filled sensor apparatus 100 loaded within the reader apparatus 190.
When the sensor apparatus 100 is positioned within the container receiving space 1306, a reference electrode contact 1308 of the reader apparatus 190 can be placed or moved into contact with the reference electrode material 149 positioned on the container cap 116 (see, e.g., Fig. 1D) of the sensor apparatus 100. Moreover, when the sensor apparatus 100 is positioned within the container receiving space 1306, an active electrode contact 1310 of the reader apparatus 190 can be placed or moved into contact with a conductive substrate layer or conductive contact (e.g., any of the conductive substrate 406 of Figs. 4B-4D or the conductive contact layer 502 of Fig. 5A) of the active sensor 106.

[0196] In some embodiments, the reference electrode contact 1308 and the active electrode contact 1310 can comprise one or more conductive pogo or spring-loaded pins, conductive leaf contacts, or a combination thereof. More specifically, the conductive pogo pins or leaf contacts can be made of copper, nickel, stainless steel, or alloys thereof.

[0197] The reference electrode contact 1308 and the active electrode contact 1310 can be electrically coupled to a signal readout control unit 1303. The signal readout control unit 1303 can comprise one or more processors, chipsets, or chip modules programmed to convert and read signals obtained from the active sensor 106 and the reference sensor 122 of the sensor apparatus 100.

[0198] Fig. 13B also illustrates that the reader apparatus 190 can comprise a thermal control module 1305 and an aeration control module 1307. The thermal control module 1305 can be configured to incubate the sample-filled sensor apparatus 100. The thermal control module 1305 can incubate the sensor apparatus 100 by heating at least part of the sensor apparatus 100 via a heating block 1318 (see, e.g., Fig. 13D). In some embodiments, the heating block 1318 can heat a lateral side of the sample chamber 108 opposite the active sensor 106. In certain embodiments, the heating block 1318 can partially surround or cradle the sample chamber 108 to heat the sensor apparatus 100.

[0199] In some embodiments, the heating block 1318 can be made in part of aluminum.
In other embodiments, the heating block 1318 can be made in part of another type of heat conducting metallic material.

[0200] The sensor apparatus 100 can be heated to an incubation temperature of between about 30 C and 40 C (e.g., about 35 C 2 C). The sensor apparatus 100 can be incubated for an incubation period. The incubation period can range from 15 minutes to over 48 hours. The incubation period can be adjusted based on the type of infectious agent suspected in the sample.

[0201] In some embodiments, the thermal control module 1305 can be controlled by the main controller 1301 (see, e.g., Fig. 13C) of the reader apparatus 190. In other embodiments, the thermal control module 1305 can be controlled by another controller or module within the reader apparatus 190 or by the signal readout control unit 1303.

[0202] In some embodiments, a nutrient solution or stimulus solution can be introduced into the sample chamber 108 before the sensor apparatus 100 is incubated. For example, the nutrient solution can be a solution containing bacto-tryptone, yeast extract, beef extract, cation-adjusted Mueller Hinton Broth (CAMHB), starch, an acid hydrolysate of casein, calcium chloride, magnesium chloride, sodium chloride, blood or lysed blood including lysed horse blood (LHB), a CAMHB-LHB mixture, glucose, or a combination thereof. The nutrient solution can be used to counteract the buffering effects of ions or substances present in the sample when the sample is composed of a bodily fluid.

[0203] The aeration control module 1307 can be configured to aerate the sample within the sample chamber 108 by pumping a gas 162 (see, e.g., Fig. 1D) into the chamber cavity 109 containing the sample. The gas 162 can be pumped into the sample chamber through an aeration port 160 defined along the bottom of the sample chamber 108 (see, e.g., Fig. 1D).

[0204] Aerating the sample can enhance a growth rate of infectious agents within the sample by increasing the supply of oxygen to such infectious agents. Moreover, aerating the sample can also enable detachment of the infectious agents from the interior walls of the sample chamber 108 so as to inhibit biofilm formation.

[0205] Fig. 13C illustrates a perspective view of a portion of the reader apparatus 190 with the reader housing 1300 removed. As shown in Fig. 13C, the aeration control module 1307 can delivery gas 162 via a gas delivery conduit 1312 connecting the aeration control module 1307 to the sensor apparatus 100. In some embodiments, at least a segment of the gas delivery conduit 1312 can be positioned along or wound around a base or bottom portion of the reader apparatus 190.

[0206] Fig. 13D illustrates a close-up view of a gas nozzle 1314 being connected to the bottom of the sensor apparatus 100 to aerate the sample within the sample chamber 108.

The gas nozzle 1314 can be disposed at a terminal or distal end of the gas delivery conduit 1312.

[0207] As shown in Fig. 13D, the gas nozzle 1314 can connect to the aeration port 160 at the bottom of the sample chamber 108 via a nozzle interface 1316. In some embodiments, the nozzle interface 1316 can be an 0-ring. In other embodiments, the nozzle interface 1316 can be another type of gasket or fluid-sealing interface.

[0208] In some embodiments, the gas 162 can be ambient air (e.g., the air in a laboratory, clinical setting, or testing facility). In other embodiments, the gas 162 can comprise a combination of pressurized oxygen, carbon dioxide, nitrogen, and argon.
Aerating the sample can accelerate the growth of a microbial population within the sample by providing an oxygen rich environment within the sample chamber 108.

[0209] The aeration control module 1307 can pump gas 162 into the sample chamber 108 at a constant flow rate of between about 1.0 mL/min and 10.0 mL/min.

[0210] In some embodiments, the aeration control module 1307 can be controlled by the main controller 1301 (see, e.g., Fig. 13C). In other embodiments, the aeration control module 1307 can be controlled by another controller or module within the reader apparatus 190 or by the signal readout control unit 1303. For example, the amount of gas 162 (e.g., ambient air) pumped or otherwise directed into the sample chamber 108 can be dictated by a change in a solution characteristic of the sample detected by the reader apparatus 190 or a lack of any such change.

[0211] Fig. 14 illustrates a method 1400 of making a sensor apparatus 100 for measuring a solution characteristic of a sample. The method 1400 can comprise cleaning a conductive substrate 406 (e.g., a sheet of stainless steel such as 316 SS) with an acid and base treatment in step 1402.

[0212] The conductive substrate 406 can first be cleaned with a series of acid and base treatments to remove any impurities or surface contaminants (e.g., free iron).
Such treatments can be performed with nitric acid (10%) followed by ammonium hydroxide (175mM), isopropyl alcohol (99%), or acetone. In other embodiments, the conductive substrate 406 can be cleaned and descaled using other acids, bases, alcohols, solvents, or other chemicals.

[0213] The method 1400 can also comprise depositing an adhesion material on one side of the cleaned conductive substrate 406 until an adhesion layer 408 forms on the cleaned conductive substrate 406 in step 1404. In some embodiments, the adhesion layer 408 can be deposited by a sputter deposition technique such as physical vapor deposition (PVD). In some embodiments, the adhesion layer 408 can be a layer of chromium (Cr).
Chromium can be selected because it creates a bond to the chromium in the stainless steel of the conductive substrate 406. In other embodiments, the adhesion layer 408 can also be a layer of gold (Au) or nickel (Ni).

[0214] Step 1404 can also comprise depositing an adhesion material (e.g., Cr, Au, or Ni) until the adhesion layer 408 is at least 20 nm thick.

[0215] The method 1400 can further comprise depositing an active electrode material on the adhesion layer 408 until an active electrode layer 132 forms on the adhesion layer 408 in step 1406. The active electrode layer 132 can be a noble metal layer such as a platinum or gold layer when the sensor apparatus 100 is to be used as an ORP
sensor.
Depositing the active electrode layer 132 can comprise depositing an active electrode material (e.g., Pt) using sputter deposition (e.g., PVD), evaporation deposition, or electrodeposition. For certain processes, such as the evaporation deposition, the conductive substrate 406 can be precleaned in vacuum with argon (Ar) plasma etching. In other embodiments, the active electrode material can be applied using ink screen-printing.

[0216] Step 1406 can also comprise depositing an active electrode material until the active electrode layer 132 is at least 50 nm thick. Step 1406 can further comprise depositing the active electrode material until the active electrode layer 132 is at least 400 nm thick. The applicants discovered that this minimum thickness is needed to prevent nano-sized holes from forming in the active electrode layer 132 that would allow fluid (e.g., the fluid sample) to make its way through the active electrode layer 132 and into contact with other layers of the active sensor 106 (thereby adversely affecting the measurement). Moreover, when an antimicrobial material (e.g., copper or nickel) is used as the conductive substrate 406, the active electrode layer 132 having a minimum thickness of 400 nm can act as a barrier to protect the microbes in the fluid sample.

[0217] In certain embodiments, the adhesion layer 408 can be deposited in a vacuum chamber and the active electrode layer 132 can be deposited subsequent to the adhesion layer 408 in the same vacuum chamber.

[0218] Alternatively, step 1406 can comprise depositing a metal layer and surface modifying the metal layer to create a metal oxide layer. For example, step 1406 can comprise depositing a platinum layer and oxidizing the platinum layer to create a platinum oxide (Pt02) layer serving as the active electrode layer 132. The active electrode layer 132 can be a metal oxide layer (e.g., platinum oxide or tantalum oxide) when the sensor apparatus 100 is to be used as a pH sensor.

[0219] The method 1400 can also comprise singulating the conductive substrate 406 covered by the adhesion layer 408 and the active electrode layer 132 in step 1408. The conductive substrate 406 covered by the adhesion layer 408 and the active electrode layer 132 can be singulated by laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or sawing. The conductive substrate 406 covered by the adhesion layer 408 and the active electrode layer 132 can be singulated to yield an active sensor 106 sized to cover a window opening 114 defined along the chamber lateral wall 112 of the sample chamber 108 (see, e.g., Figs. 1B-1D and 2).

[0220] The method 1400 can further comprise coupling the active sensor 106 to at least part of the chamber lateral wall 112 in step 1410. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that no part of the active sensor 106 extends into a chamber cavity 109 within the sample chamber 108 and the active electrode layer 132 faces the chamber cavity 109 to allow any sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that the active sensor 106 (including the active electrode layer 132) is positioned radially outward from an interior-facing or cavity-facing side of the chamber lateral wall 112 and the lateral sides 136 of the active sensor 106 are not in fluid communication with the chamber cavity 109.

[0221] In some embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can further comprise applying a bead of adhesive 138 to a part of the chamber lateral wall 112 within a recessed portion 134 defined along the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the bead of adhesive 138 within the recessed portion 134, and curing the adhesive 138.

[0222] In alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise insert-molding the active sensor 106 into the chamber lateral wall 112 while the sample chamber 108 is formed by injection molding.

[0223] In further alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise focally melting (e.g., by ultrasonic welding) a part of the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the melted part of the chamber lateral wall 112, and allowing the melted part of the chamber lateral wall 112 to cool to affix the active sensor 106 to the chamber lateral wall 112.

[0224] Fig. 15 illustrates yet another method 1500 of making a sensor apparatus 100 for measuring a solution characteristic of a sample. The method 1500 can comprise providing a non-conductive printed circuit board (PCB) substrate 500 (see, e.g., Fig. 5A) in step 1502.

[0225] The method 1500 can also comprise depositing an active electrode material on one side of the non-conductive PCB substrate 500 until an active electrode layer 132 forms on the non-conductive PCB substrate 500 in step 1504. Step 1504 can also comprise depositing an active electrode material until the active electrode layer 132 is at least 50 nm thick. Step 1504 can further comprise depositing an active electrode material until the active electrode layer 132 is at least 400 nm thick. After the deposition step, the active electrode layer 132 can be electrically coupled to conductive contacts or a conductive contact layer 502 of the non-conductive PCB substrate 500 by conductive vias extending through the non-conductive PCB substrate 500.

[0226] The active electrode layer 132 can be a noble metal layer such as a platinum or gold layer when the sensor apparatus 100 is to be used as an ORP sensor.
Depositing the active electrode layer 132 can comprise depositing an active electrode material (e.g., Pt) using sputter deposition (e.g., PVD), evaporation deposition, or electrodeposition.

[0227] The method 1500 can also comprise singulating the non-conductive PCB

substrate 500 covered by the active electrode layer 132 to yield an active sensor 106 sized to cover a window opening 114 defined along a chamber lateral wall 112 of a sample chamber 108 in step 1506. The non-conductive PCB substrate 500 covered by the active electrode layer 132 can be singulated by laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or sawing. The active sensor 106 can comprise at least one conductive via 504 extending through the PCB substrate 500.

[0228] The method 1500 can further comprise coupling the active sensor 106 to at least part of the chamber lateral wall 112 in step 1508. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that no part of the active sensor 106 extends into a chamber cavity 109 within the sample chamber 108 and the active electrode layer 132 faces the chamber cavity 109 to allow any sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that the active sensor 106 (including the active electrode layer 132) is positioned radially outward from an interior-facing or cavity-facing side of the chamber lateral wall 112 and the lateral sides 136 of the active sensor 106 are not in fluid communication with the chamber cavity 109.

[0229] In some embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can further comprise applying a bead of adhesive 138 to a part of the chamber lateral wall 112 within a recessed portion 134 defined along the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the bead of adhesive 138 within the recessed portion 134, and curing the adhesive 138.

[0230] In alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise insert-molding the active sensor 106 into the chamber lateral wall 112 while the sample chamber 108 is formed by injection molding.

[0231] In further alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise focally melting (e.g., by ultrasonic welding) a part of the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the melted part of the chamber lateral wall 112, and allowing the melted part of the chamber lateral wall 112 to cool to affix the active sensor 106 to the chamber lateral wall 112.

[0232] Fig. 16 illustrates yet another method 1600 of making a sensor apparatus 100 for measuring a solution characteristic of a sample. The method 1600 can comprise providing a non-conductive polymeric substrate 700 comprising a plurality of through-holes 702 in step 1602. The method 1600 can also comprise depositing a conductive contact layer 502 on one side of the polymeric substrate 700 in step 1604.
Depositing the conductive contact layer 502 can comprise depositing an electrically conductive material (e.g., Au) on the polymeric substrate 700 using sputter deposition (e.g., PVD), evaporation deposition, or electrodeposition.

[0233] The method 1600 can further comprise depositing an active electrode layer 132 on another side of the polymeric substrate 700 in step 1606. Depositing the active electrode layer 132 can comprise depositing an active electrode material (e.g., Pt) on the polymeric substrate 700 using sputter deposition (e.g., PVD), evaporation deposition, or electrodeposition.

[0234] One end of each of the through-holes 702 can be covered by the active electrode layer 132 and the other end of each of the through-holes 702 can be covered by the conductive contact layer 502. The active electrode layer 132, after the deposition steps, can be electrically coupled to the conductive contact layer 502 via a conductive coating covering lateral sides of the through-holes 702.

[0235] The method 1600 can also comprise singulating the non-conductive polymeric substrate 700 covered by the active electrode layer 132 and the conductive contact layer 502 to yield an active sensor 106 sized to cover a window opening 114 defined along a chamber lateral wall 112 of a sample chamber 108 in step 1608. The non-conductive polymeric substrate 700 covered by the active electrode layer 132 and the conductive contact layer 502 can be singulated by laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or sawing. The active sensor 106 can comprise at least one through-hole 702 extending through the non-conductive polymeric substrate 700.

[0236] The method 1600 can further comprise coupling the active sensor 106 to at least part of the chamber lateral wall 112 in step 1610. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that no part of the active sensor 106 extends into a chamber cavity 109 within the sample chamber 108 and the active electrode layer 132 faces the chamber cavity 109 to allow any sample within the chamber cavity 109 to be in fluid contact with the active electrode layer 132 through at least part of the chamber lateral wall 112 surrounding the window opening 114. The active sensor 106 can be coupled to at least part of the chamber lateral wall 112 such that the active sensor 106 (including the active electrode layer 132) is positioned radially outward from an interior-facing or cavity-facing side of the chamber lateral wall 112 and the lateral sides 136 of the active sensor 106 are not in fluid communication with the chamber cavity 109.

[0237] In some embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can further comprise applying a bead of adhesive 138 to a part of the chamber lateral wall 112 within a recessed portion 134 defined along the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the bead of adhesive 138 within the recessed portion 134, and curing the adhesive 138.

[0238] In alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise insert-molding the active sensor 106 into the chamber lateral wall 112 while the sample chamber 108 is formed by injection molding.

[0239] In further alternative embodiments, coupling the active sensor to at least part of the chamber lateral wall 112 of the sample chamber 108 can comprise focally melting (e.g., by ultrasonic welding) a part of the chamber lateral wall 112 surrounding the window opening 114, pressing or placing the active sensor 106 onto the melted part of the chamber lateral wall 112, and allowing the melted part of the chamber lateral wall 112 to cool to affix the active sensor 106 to the chamber lateral wall 112.

[0240] A number of embodiments have been described. Nevertheless, it will be understood by one of ordinary skill in the art that various changes and modifications can be made to this disclosure without departing from the spirit and scope of the embodiments.
Elements of systems, devices, apparatus, and methods shown with any embodiment are exemplary for the specific embodiment and can be used in combination or otherwise on other embodiments within this disclosure. For example, the steps of any methods depicted in the figures or described in this disclosure do not require the particular order or sequential order shown or described to achieve the desired results. In addition, other steps operations may be provided, or steps or operations may be eliminated or omitted from the described methods or processes to achieve the desired results. Moreover, any components or parts of any apparatus or systems described in this disclosure or depicted in the figures may be removed, eliminated, or omitted to achieve the desired results. In addition, certain components or parts of the systems, devices, or apparatus shown or described herein have been omitted for the sake of succinctness and clarity.

[0241] Accordingly, other embodiments are within the scope of the following claims and the specification and/or drawings may be regarded in an illustrative rather than a restrictive sense.

[0242] Each of the individual variations or embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other variations or embodiments.
Modifications may be made to adapt a particular situation, material, composition of matter, process, process act(s) or step(s) to the objective(s), spirit or scope of the present invention.

[0243] Methods recited herein may be carried out in any order of the recited events that is logically possible, as well as the recited order of events. Moreover, additional steps or operations may be provided or steps or operations may be eliminated to achieve the desired result.

[0244] Furthermore, where a range of values is provided, every intervening value between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the invention. Also, any optional feature of the inventive variations described may be set forth and claimed independently, or in combination with any one or more of the features described herein. For example, a description of a range from 1 to 5 should be considered to have disclosed subranges such as from 1 to 3, from 1 to 4, from 2 to 4, from 2 to 5, from 3 to 5, etc. as well as individual numbers within that range, for example 1.5, 2.5, etc. and any whole or partial increments therebetween.

[0245] All existing subject matter mentioned herein (e.g., publications, patents, patent applications) is incorporated by reference herein in its entirety except insofar as the subject matter may conflict with that of the present invention (in which case what is present herein shall prevail). The referenced items are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such material by virtue of prior invention.

[0246] Reference to a singular item, includes the possibility that there are plural of the same items present. More specifically, as used herein and in the appended claims, the singular forms "a," "an," "said" and "the" include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as "solely," "only" and the like in connection with the recitation of claim elements, or use of a "negative" limitation. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0247] In understanding the scope of the present disclosure, the term "comprising" and its derivatives, as used herein, are intended to be open-ended terms that specify the presence of the stated features, elements, components, groups, integers, and/or steps, but do not exclude the presence of other unstated features, elements, components, groups, integers and/or steps. The foregoing also applies to words having similar meanings such as the terms, "including", "having" and their derivatives. Also, the terms "part,"
"section,"
"portion," "member" "element," or "component" when used in the singular can have the dual meaning of a single part or a plurality of parts. As used herein, the following directional terms "forward, rearward, above, downward, vertical, horizontal, below, transverse, laterally, and vertically" as well as any other similar directional terms refer to those positions of a device or piece of equipment or those directions of the device or piece of equipment being translated or moved. Finally, terms of degree such as "substantially", "about" and "approximately" as used herein mean a reasonable amount of deviation (e.g., a deviation of up to 0.1%, 1%, 5%, or 10%, as such variations are appropriate) from the specified value such that the end result is not significantly or materially changed.

[0248] This disclosure is not intended to be limited to the scope of the particular forms set forth, but is intended to cover alternatives, modifications, and equivalents of the variations or embodiments described herein. Further, the scope of the disclosure fully encompasses other variations or embodiments that may become obvious to those skilled in the art in view of this disclosure.

Claims

What is claimed is:

1. A sensor apparatus for measuring a solution characteristic of a sample, the sensor apparatus comprising:
a sample container comprising a sample chamber, wherein the sample chamber comprises a chamber lateral wall surrounding a chamber cavity configured to receive the sample, a reference sensor comprising a reference electrode material and a wick in fluid communication with the sample chamber such that at least some of the sample is drawn by the wick in a direction of the reference electrode material; and an active sensor made of a conductive substrate covered in part by an active electrode layer, wherein the active sensor is coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall, wherein no part of the active sensor extends into the chamber cavity, and wherein the active electrode layer faces the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening, wherein the solution characteristic of the sample is determined based on a potential difference measured between the active sensor and the reference sensor when the reference sensor and the active sensor are electrically coupled to a reader.

2. The sensor apparatus of claim 1, wherein the solution characteristic measured is an oxidation reduction potential and wherein the active electrode layer is a platinum layer.

3. The sensor apparatus of claim 1, wherein the solution characteristic measured is pH and wherein the active electrode layer comprises a platinum oxide layer and a tantalum oxide layer.

4. The sensor apparatus of claim 1, wherein the conductive substrate is stainless steel.

5. The sensor apparatus of claim 1, wherein the active electrode layer has an active electrode layer thickness of between about 50 nm and 500 nm.

6. The sensor apparatus of claim 5, wherein the active sensor further comprises an adhesion layer in between the conductive substrate and the active electrode layer, wherein the adhesion layer has an adhesion layer thickness of between 5 nm and 50 nm.

7. The sensor apparatus of claim 6, wherein a ratio of the adhesion layer thickness to the active electrode layer thickness is between about 1:10 and 1:20.

8. The sensor apparatus of claim 6, wherein the adhesion layer is a chromium layer.

9. The sensor apparatus of claim 1, wherein the active electrode layer has an active electrode layer thickness of at least 50 nm.

10. The sensor apparatus of claim 1, wherein the active sensor is insert-molded into the chamber lateral wall while the sample container is formed by injection molding.

11. The sensor apparatus of claim 1, wherein the active sensor is press-molded into the chamber lateral wall after the sample container is formed by injection molding.

12. The sensor apparatus of claim 1, wherein the chamber lateral wall comprises a recessed portion surrounding the window opening and defined along an exterior side of the chamber lateral wall and wherein the active sensor is adhered to at least part of the chamber lateral wall within the recessed portion via an adhesive.

13. The sensor apparatus of claim 1, wherein the active sensor comprises an active electrode side, a conductive substrate side opposite the active electrode side, and lateral sides, wherein the lateral sides are covered by at least one of the chamber lateral wall and an adhesive to prevent the lateral sides from contacting the sample.

14. The sensor apparatus of claim 1, wherein the sample chamber is made in part of at least one of polyoxymethylene, polyamide, polyethylene, acrylonitrile butadiene styrene, polycarbonate, and polypropylene.

15. The sensor apparatus of claim 1, wherein the reference electrode material is a cured or hardened silver-silver chloride ink deposited or otherwise applied on a wick proximal end of the wick.

16. A sensor apparatus for measuring a solution characteristic of a sample, the sensor apparatus comprising:
a sample container comprising a sample chamber, wherein the sample chamber comprises a chamber lateral wall surrounding a chamber cavity configured to receive the sample, a reference sensor comprising a reference electrode material and a wick in fluid communication with the sample chamber such that at least some of the sample is drawn by the wick in a direction of the reference electrode material; and an active sensor made of a non-conductive printed circuit board (PCB) substrate covered in part by an active electrode layer, wherein the active electrode layer is electrically coupled to conductive contacts of the PCB substrate by a conductive via extending through the PCB substrate, wherein the active sensor is coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall, wherein no part of the active sensor extends into the chamber cavity, and wherein the active electrode layer faces the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening, wherein the solution characteristic of the sample is determined based on a potential difference measured between the active sensor and the reference sensor when the reference sensor and the active sensor are electrically coupled to a reader.

17. The sensor apparatus of claim 16, wherein the PCB substrate is a flexible PCB
substrate.

18. The sensor apparatus of claim 16, wherein the PCB substrate is made in part of polyimide.

19. The sensor apparatus of claim 16, wherein the PCB substrate is made in part of an FR-4 composite material.

20. The sensor apparatus of claim 16, wherein the conductive via is made of copper.

21. The sensor apparatus of claim 16, wherein the solution characteristic measured is an oxidation reduction potential and wherein the active electrode layer is a platinum layer or a gold layer.

22. The sensor apparatus of claim 16, wherein the active electrode layer has an active electrode layer thickness of at least 50 nm.

23. The sensor apparatus of claim 16, wherein the active sensor is insert-molded into the chamber lateral wall while the sample container is formed by injection molding.

24. The sensor apparatus of claim 16, wherein the solution characteristic measured is pH
and wherein the active electrode layer is a metal oxide layer.

25. The sensor apparatus of claim 16, wherein the chamber lateral wall comprises a recessed portion surrounding the window opening and defined along an exterior side of the chamber lateral wall and wherein the active sensor is adhered to at least part of the chamber lateral wall within the recessed portion via an adhesive.

26. The sensor apparatus of claim 16, wherein the active sensor comprises an active electrode side covered by the active electrode layer, a conductive contact side opposite the active electrode side and comprising the conductive contacts, and lateral sides, wherein the lateral sides are covered by at least one of the chamber lateral wall and an adhesive to prevent the lateral sides from contacting the sample.

27. A sensor apparatus for measuring a solution characteristic of a sample, the sensor apparatus comprising:
a sample container comprising a sample chamber, wherein the sample chamber comprises a chamber lateral wall surrounding a chamber cavity configured to receive the sample, a reference sensor comprising a reference electrode material and a wicking component in fluid communication with the sample chamber such that at least some of the sample is drawn by the wicking component in a direction of the reference electrode material; and an active sensor made of a non-conductive polymeric substrate comprising a through-hole, wherein one side of the polymeric substrate and one end of the through-hole is covered by an active electrode layer and wherein another side of the polymeric substrate and the other end of the through-hole is covered by a conductive layer, wherein the active electrode layer is electrically coupled to the conductive layer via a conductive coating covering lateral sides of the through-hole, wherein the active sensor is coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall, wherein no part of the active sensor extends into the chamber cavity, and wherein the active electrode layer faces the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening, wherein the solution characteristic of the sample is determined based on a potential difference measured between the active sensor and the reference sensor when the reference sensor and the active sensor are electrically coupled to a reader.

28. The sensor apparatus of claim 27, wherein the active electrode layer is a platinum layer.

29. The sensor apparatus of claim 27, wherein the conductive layer is a gold layer.

30. The sensor apparatus of claim 27, wherein the active electrode layer has an active electrode layer thickness of at least 50 nm.

31. The sensor apparatus of claim 27, wherein the active sensor is a rectangular piece having a width dimension of between about 100 um and 6.0 mm and a length dimension of between about 100 um and 6.0 mm.

32. The sensor apparatus of claim 27, wherein the through-hole has a diameter between about 10 um to 100 um.

33. The sensor apparatus of claim 27, wherein the conductive coating covering the lateral sides of the through-hole is a coating of platinum.

34. The sensor apparatus of claim 27, wherein the conductive coating covering the lateral sides of the through-hole is a coating of gold.

35. The sensor apparatus of claim 27, wherein the active sensor is insert-molded into the chamber lateral wall while the sample container is formed by injection molding.

36. The sensor apparatus of claim 27, wherein the active sensor is press-molded into the chamber lateral wall after the sample container is formed by injection molding.

37. The sensor apparatus of claim 27, wherein the chamber lateral wall comprises a recessed portion surrounding the window opening and defined along an exterior side of the chamber lateral wall and wherein the active sensor is adhered to at least part of the chamber lateral wall within the recessed portion via an adhesive.

38. A sensor apparatus for measuring a solution characteristic of a sample, the sensor apparatus comprising:
a sample container comprising a sample chamber, wherein the sample chamber comprises a chamber lateral wall surrounding a chamber cavity configured to receive the sample, a reference sensor comprising a reference electrode material and a wicking component in fluid communication with the sample chamber such that at least some of the sample is drawn by the wicking component in a direction of the reference electrode material; and an active sensor made of a conductive dowel covered in part by an active electrode layer, wherein the active sensor is coupled to at least part of the chamber lateral wall at a window opening defined along the chamber lateral wall, wherein the part of the conductive dowel covered by the active electrode layer extends into the chamber cavity to allow the sample within the chamber cavity to be in fluid contact with the active electrode layer, and wherein an end of the conductive dowel not extending into the chamber cavity extends out of the chamber lateral wall, wherein the solution characteristic of the sample is determined based on a potential difference measured between the active sensor and the reference sensor when the reference sensor and the active sensor are electrically coupled to a reader.

39. The sensor apparatus of claim 38, wherein the solution characteristic measured is an oxidation reduction potential and wherein the active electrode layer is a platinum layer.

40. The sensor apparatus of claim 38, wherein the conductive dowel is made in part of stainless steel.

41. The sensor apparatus of claim 38, wherein the active sensor is insert-molded into the chamber lateral wall while the sample container is formed by injection molding.

42. The sensor apparatus of claim 38, wherein the active sensor is adhered to parts of the chamber lateral wall surrounding the window opening.

43. The sensor apparatus of claim 38, wherein the conductive dowel is shaped substantially as a cylinder having rounded edges.

44. The sensor apparatus of claim 38, wherein the active electrode layer has an active electrode layer thickness of at least 50 nm.

45. A method of making a sensor apparatus for measuring a solution characteristic of a sample, the method comprising:
cleaning a conductive substrate with an acid and base treatment;
depositing an adhesion layer on one side of the conductive substrate;
depositing an active electrode layer on the adhesion layer;
singulating the conductive substrate covered by the adhesion layer and the active electrode layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber; and coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

46. The method of claim 45, wherein cleaning the conductive substrate further comprises treating the conductive substrate with nitric acid followed by treating the conductive substrate with ammonium hydroxide, isopropyl alcohol, or acetone.

47. The method of claim 45, wherein singulating the conductive substrate further comprises laser cutting, metal shearing, hot wire cutting, dye cutting, stamping, or sawing the conductive substrate.

48. The method of claim 45, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
applying a bead of adhesive to a part of the chamber lateral wall within a recessed portion defined along the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the bead of adhesive within the recessed portion;
and curing the adhesive.

49. The method of claim 45, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises insert-molding the active sensor into the chamber lateral wall while the sample chamber is formed by injection molding.

50. The method of claim 45, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
focally melting a part of the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the melted part of the chamber lateral wall;
and allowing the melted part of the chamber lateral wall to cool to affix the active sensor to the chamber lateral wall.

51. The method of claim 45, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer until a thickness of the active electrode layer is at least 50 nm.

52. The method of claim 51, wherein the active electrode material is platinum when the solution characteristic measured is an oxidation reduction potential (ORP) of the sample.

53. The method of claim 45, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer using sputter deposition, evaporation deposition, electrodeposition, or ink screen-printing.

54. The method of claim 53, wherein depositing the adhesion layer comprises depositing an adhesion material making up the adhesion layer using sputter deposition.

55. The method of claim 54, wherein the adhesion layer is deposited first in a vacuum chamber and the active electrode layer is deposited subsequently in the vacuum chamber.

56. The method of claim 51, wherein the active electrode material is a metal oxide when the solution characteristic measured is a pH of the sample.

57. The method of claim 56, wherein the metal oxide is platinum oxide and the platinum oxide covers a platinum layer deposited on the adhesion layer.

58. A method of making a sensor apparatus for measuring a solution characteristic of a sample, the method comprising:
providing a non-conductive printed circuit board (PCB) substrate;
depositing an active electrode layer on one side of the PCB substrate, wherein the active electrode layer, after the deposition step, is electrically coupled to conductive contacts of the PCB substrate by conductive vias extending through the PCB substrate;
singulating the PCB substrate covered by the active electrode layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber, and wherein the active sensor comprises at least one conductive via extending through the PCB substrate; and coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

59. The method of claim 58, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer using sputter deposition, evaporation deposition, and electrodeposition.

60. The method of claim 58, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer until a thickness of the active electrode layer is at least 50 nm.

61. The method of claim 59, wherein the active electrode material is platinum or gold when the solution characteristic measured is an oxidation reduction potential (ORP) of the sample.

62. The method of claim 61, wherein the conductive contacts are made in part of gold.

63. The method of claim 58, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
applying a bead of adhesive to a part of the chamber lateral wall within a recessed portion defined along the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the bead of adhesive within the recessed portion;
and curing the adhesive.

64. The method of claim 58, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises insert-molding the active sensor into the chamber lateral wall while the sample chamber is formed by injection molding.

65. The method of claim 58, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
focally melting a part of the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the melted part of the chamber lateral wall;
and allowing the melted part of the chamber lateral wall to cool to affix the active sensor to the chamber lateral wall.

66. A method of making a sensor apparatus for measuring a solution characteristic of a sample, the method comprising:
providing a non-conductive polymeric substrate comprising a plurality of through-holes;
depositing a conductive layer on one side of the polymeric substrate;
depositing an active electrode layer on another side of the polymeric substrate, wherein one end of the through-holes are covered by the active electrode layer and the other end of the through-holes are covered by the conductive layer, wherein the active electrode layer, after the deposition steps, is electrically coupled to the conductive layer via a conductive coating covering lateral sides of the through-holes;
singulating the polymeric substrate covered by the active electrode layer and the conductive layer to yield an active sensor sized to cover a window opening defined along a chamber lateral wall of a sample chamber, wherein the active sensor comprises at least one through-hole covered by the active electrode layer and the conductive layer; and coupling the active sensor to at least part of the chamber lateral wall such that no part of the active sensor extends into a chamber cavity within the sample chamber and the active electrode layer faces the chamber cavity to allow any sample within the chamber cavity to be in fluid contact with the active electrode layer through at least part of the chamber lateral wall surrounding the window opening.

67. The method of claim 66, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer using sputter deposition, evaporation deposition, and electrodeposition.

68. The method of claim 66, wherein depositing the active electrode layer comprises depositing an active electrode material making up the active electrode layer until a thickness of the active electrode layer is at least 50 nm.

69. The method of claim 67, wherein the active electrode material is platinum when the solution characteristic measured is an oxidation reduction potential (ORP) of the sample.

70. The method of claim 66, wherein depositing the conductive comprises depositing a conductive material on the other side of the polymeric substrate.

71. The method of claim 70, wherein the conductive material is gold.

72. The method of claim 66, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
applying a bead of adhesive to a part of the chamber lateral wall within a recessed portion defined along the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the bead of adhesive within the recessed portion;
and curing the adhesive.

73. The method of claim 66, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises insert-molding the active sensor into the chamber lateral wall while the sample chamber is formed by injection molding.

74. The method of claim 66, wherein coupling the active sensor to at least part of the chamber lateral wall of the sample chamber further comprises:
focally melting a part of the chamber lateral wall surrounding the window opening;
pressing the active sensor onto the melted part of the chamber lateral wall;
and allowing the melted part of the chamber lateral wall to cool to affix the active sensor to the chamber lateral wall.