Device and method for checking influence of drugs on microorganisms

Abstract

Devices, systems, and methods for performing biological and chemical assays are provided.

Description

[0001] Cross-references

[0002] This application is a divisional application of the Chinese national phase application with application number 2018800638305 and filing date August 1, 2018. Like the parent application, this application claims U.S. Provisional Patent Application No. 62 / 539,677, filed on August 1, 2017, which is incorporated herein by reference in its entirety for all purposes. Technical Field

[0003] The present invention particularly relates to apparatus and methods for performing biological and chemical assays, especially microbiological assays. Background Art

[0004] In diagnosis and clinical setting, often need to check the influence of medicine on microorganism.For example, usually use susceptibility test to help health care personnel determine which medicine or treatment strategy is most effective in treating patient infection, susceptibility test determines which antimicrobial will suppress the growth of the bacterium or fungus that causes patient-specific infection.Yet, in fact, this test is normally laborious, time consuming, and needs complicated laboratory environment and professional processing.Especially, be generally related to the culture of cultivating infection site and the conventional method of monitoring culture growth in time after using antimicrobial, the time that spends when facing frequent, rapid development or even life-threatening infection is too long.Therefore, what this area is highly recognized is, need quick, easy to handle and accurate alternative test. Summary of the Invention

[0005] The present invention provides an apparatus and method for solving the above-mentioned problems in examining the effects of drugs on microorganisms.

[0006] One aspect of the invention uses (a) two plates to compress a sample containing microorganisms to be examined into a thin layer, and (b), following (a), uses an imaging process to count and / or track the microorganisms over a period of time or at discrete time points.

[0007] Another aspect of the present invention uses spacers to control the final sample thickness, thereby aiding in determining microbial concentration.

[0008] Another aspect of the present invention provides coating the drug to be examined on one or both plates, since the coated drug is capable of dissolving and diffusing in the sample when in contact with the sample.

[0009] Another aspect of the present invention provides a multiplex capability for drug effect testing using two plates and drugs coated at different locations on the plates.

[0010] Another aspect of the present invention uses a viability dye that is added to the sample when the sample is loaded between two provided plates and the microbial activity is monitored with the aid of the dye.

[0011] Another aspect of the present invention provides the use of a mobile communication device (eg, a cell phone) for imaging microorganisms loaded between two provided plates.

[0012] Yet another aspect of the present invention uses mobile communication devices, in addition to imaging, to notify medical personnel or other parties of test results. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Those skilled in the art will appreciate that the drawings described below are for illustrative purposes only. The drawings are not intended to limit the scope of the present invention in any way. Some of the drawings are not drawn to scale. In the drawings that provide experimental data points, the lines connecting the data points are used only to guide the observation of the data and have no other purpose.

[0014] Figure 1 Shown is an example of a generalized QMAX (Q: quantization); M: amplification; A: addition of reagents; X: acceleration; also known as a compression regulated open flow (CROF) device.

[0015] Figure 2 An exemplary embodiment of the present invention is shown in which a QMAX device is used to monitor microorganisms.

[0016] Figure 3 An exemplary embodiment of the invention is shown, wherein the proliferation of monitored microorganisms between two plates.

[0017] Figure 4 Another exemplary embodiment of the present invention is shown wherein the test agent affects the survival of a microorganism.

[0018] Detailed Description of Exemplary Embodiments

[0019] The following detailed description illustrates some embodiments of the present invention by way of example and not limitation. The section headings and any subtitles used herein, if any, are for organizational purposes only and should not be construed as limiting the subject matter described in any way. The content under a section heading and / or subheading is not limited to the section heading and / or subheading, but applies to the entire description of the present invention.

[0020] The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present claims are not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed.

[0021] Among other things, the present invention provides apparatus, systems, and methods for performing biological and chemical assays using QMAX cards.

[0022] The exemplary embodiments disclosed herein can be combined with biological / chemical devices and assays, including but not limited to those disclosed, described, and / or referred to in the following applications:

[0023] PCT application No. PCT / US2016 / 045437, filed on August 10, 2016,

[0024] PCT application No. PCT / US2016 / 051775, filed September 14, 2016,

[0025] PCT application No. PCT / US2016 / 051794, filed September 14, 2016,

[0026] U.S. Provisional Application No. 62 / 369,181, filed July 31, 2016,

[0027] U.S. Provisional Application No. 62 / 412,006, filed October 24, 2016,

[0028] U.S. Provisional Application No. 62 / 437,339, filed December 21, 2016,

[0029] U.S. Provisional Application No. 62 / 431,639, filed December 9, 2016,

[0030] U.S. Provisional Application No. 62 / 456,065, filed February 7, 2017,

[0031] U.S. Provisional Application No. 62 / 456,488, filed February 8, 2017,

[0032] U.S. Provisional Application No. 62 / 456,287, filed February 8, 2017,

[0033] U.S. Provisional Application No. 62 / 456,528, filed February 8, 2017,

[0034] U.S. Provisional Application No. 62 / 456,537, filed February 8, 2017,

[0035] U.S. Provisional Application No. 62 / 456,612, filed February 8, 2017,

[0036] U.S. Provisional Application No. 62 / 456,631, filed February 8, 2017,

[0037] U.S. Provisional Application No. 62 / 456,596, filed February 8, 2017,

[0038] U.S. Provisional Application No. 62 / 456,590, filed February 8, 2017,

[0039] U.S. Provisional Application No. 62 / 456,638, filed February 8, 2017,

[0040] U.S. Provisional Application No. 62 / 456,598, filed February 8, 2017,

[0041] U.S. Provisional Application No. 62 / 456,552, filed February 8, 2017,

[0042] U.S. Provisional Application No. 62 / 456,603, filed February 8, 2017,

[0043] U.S. Provisional Application No. 62 / 456,585, filed February 8, 2017,

[0044] U.S. Provisional Application No. 62 / 456,628, filed February 8, 2017,

[0045] U.S. Provisional Application No. 62 / 456,504, filed February 8, 2017,

[0046] U.S. Provisional Application No. 62 / 456,988, filed February 9, 2017,

[0047] U.S. Provisional Application No. 62 / 457,084, filed February 9, 2017,

[0048] U.S. Provisional Application No. 62 / 457,031, filed February 9, 2017,

[0049] U.S. Provisional Application No. 62 / 456,904, filed February 9, 2017,

[0050] U.S. Provisional Application No. 62 / 457,075, filed February 9, 2017,

[0051] U.S. Provisional Application No. 62 / 457,009, filed February 9, 2017,

[0052] U.S. Provisional Application No. 62 / 457,133, filed February 9, 2017,

[0053] U.S. Provisional Application No. 62 / 457,103, filed February 9, 2017,

[0054] U.S. Provisional Application No. 62 / 459,267, filed February 15, 2017,

[0055] U.S. Provisional Application No. 62 / 459,303, filed February 15, 2017,

[0056] U.S. Provisional Application No. 62 / 459,337, filed February 15, 2017,

[0057] U.S. Provisional Application No. 62 / 459,232, filed February 15, 2017,

[0058] U.S. Provisional Application No. 62 / 459,160, filed February 15, 2017,

[0059] U.S. Provisional Application No. 62 / 459,972, filed February 16, 2017,

[0060] U.S. Provisional Application No. 62 / 394,753, filed September 15, 2016,

[0061] U.S. Provisional Application No. 62 / 459,496, filed February 15, 2017,

[0062] U.S. Provisional Application No. 62 / 459,554, filed February 15, 2017,

[0063] U.S. Provisional Application No. 62 / 460,047, filed February 16, 2017,

[0064] U.S. Provisional Application No. 62 / 459,598, filed February 15, 2017,

[0065] U.S. Provisional Application No. 62 / 460,083, filed February 16, 2017,

[0066] U.S. Provisional Application No. 62 / 460,076, filed February 16, 2017,

[0067] U.S. Provisional Application No. 62 / 460,062, filed February 16, 2017,

[0068] U.S. Provisional Application No. 62 / 459,920, filed February 16, 2016,

[0069] U.S. Provisional Application No. 62 / 459,577, filed February 15, 2017,

[0070] U.S. Provisional Application No. 62 / 459,602, filed February 15, 2017,

[0071] U.S. Provisional Application No. 62 / 460,069, filed February 16, 2017,

[0072] U.S. Provisional Application No. 62 / 460,088, filed February 16, 2017,

[0073] U.S. Provisional Application No. 62 / 460,091, filed February 16, 2017,

[0074] U.S. Provisional Application No. 62 / 460,757, filed February 18, 2017,

[0075] U.S. Provisional Application No. 62 / 463,578, filed February 24, 2017,

[0076] The entire contents of these applications are incorporated herein by reference.

[0077] The embodiments of these applications incorporated herein may be considered in combination with one another or as a single invention, rather than as discrete and independent documents. In addition, the exemplary embodiments disclosed herein are applicable to embodiments including, but not limited to, biological / chemical assays, QMAX cards and systems, QMAX with hinges, notches, grooved edges, and sliders, assays and devices with uniform sample thickness, smartphone detection systems, cloud computing designs, various detection methods, labels, capture and detection agents, analytes, diseases, applications, and samples; various embodiments are disclosed, described, and / or referenced in the aforementioned applications, all of which are hereby incorporated by reference in their entirety.

[0078] As used herein, the term "microorganism" refers to any organism that is small and can only be observed through a microscope, typically with an average size of less than 1 mm. Microorganisms as used herein include, but are not limited to, bacteria, fungi, archaea, viruses, protozoa, and microfauna (e.g., myxozoa, arthropods, crustaceans, and microscopic nematodes).

[0079] As used herein, the terms "drug" and "test agent" are interchangeable and refer to a biological / chemical agent whose effect on one or more microorganisms is to be examined. The term "antimicrobial agent" or "antimicrobial agent" refers to a biological / chemical agent that is capable of inhibiting the survival or growth of one or more microorganisms.

[0080] As used herein, the terms "proliferation" and "division" are interchangeable with respect to a microorganism as a subject, and refer to the action / process by which a microorganism produces new individuals by dividing a parent individual cell into two or more daughter cells, resulting in an increase in the number of microorganism individuals in the subject.

[0081] QMAX device

[0082] Figure 1An embodiment of a universal QMAX (Q: quantification; M: amplification; A: reagent addition; X: acceleration) device is shown. The universal QMAX device comprises a first plate 10 and a second plate 20. Specifically, Figure (A) shows a perspective view of the first plate 10 and the second plate 20, wherein the first plate has a spacer. However, it should be noted that the spacer can also be affixed to the second plate 20 (not shown) or to both the first plate 10 and the second plate 20 (not shown). Figure (B) shows a perspective view and a cross-sectional view of a sample 90 deposited on the first plate 10 in an open configuration. However, it should be noted that the sample 90 can also be deposited on the second plate 20 (not shown) or on both the first plate 10 and the second plate 20 (not shown). Figure (C) illustrates (i) the use of the first and second plates 10, 20 to distribute the sample 90 (the sample flows between the inner surfaces of the plates) and reduce sample thickness, and (ii) the use of the spacer and plates to adjust sample thickness in the closed configuration of the QMAX device. The inner surface of each plate has one or more binding sites and / or storage sites (not shown).

[0083] In some embodiments, the spacers 40 have a predetermined uniform height and a predetermined uniform spacer spacing. Figure 1 As shown in FIG. 4(C), the spacing between the plates and, therefore, the thickness of the sample 90 is adjusted by the spacer 40. In some embodiments, the uniform thickness of the sample 90 is substantially similar to the uniform height of the spacer 40. It should be noted that, although Figure 1 A spacer 40 is shown fixed to one of the plates, but in some embodiments, the spacer is not fixed. For example, in some embodiments, the spacer is mixed with the sample so that when the sample is compressed into a thin layer, the spacer, which acts as a rigid bead or particle of uniform size, adjusts the thickness of the sample layer.

[0084] Microbial monitoring

[0085] Conventional antimicrobial agents affect microorganisms such as bacteria or fungi by inhibiting cell proliferation processes (e.g., "bacteriostasis") or by directly killing cells (e.g., "bactericidal") by processes such as destroying the integrity of the cell wall or cell membrane. For example, penicillins and cephalosporins have a bactericidal effect by targeting the bacterial cell wall, while tetracyclines are protein synthesis inhibitors that block bacterial division. Other agents can affect microorganisms through similar mechanisms or in the opposite direction, promoting the activity and / or proliferation of microorganisms. Detecting the effect of a drug on a microorganism primarily involves monitoring one or more aspects of the microorganism after administration of the test agent, such as, but not limited to, the number of microorganisms caused by cell proliferation or cell death, proliferation rate, cell motility, cell morphology, cell membrane permeability, and cell respiration.

[0086] One aspect of the present invention is the use of a QMAX device to monitor microorganisms by one or more methods. Figure 2 An exemplary embodiment of the present invention is shown in which a QMAX device is used to monitor microorganisms. A perspective view, a top view, and a cross-sectional view taken along line aa' of the QMAX device with a deposited sample are shown, respectively. The QMAX device comprises a first plate 10, a second plate 20, and a spacer, and is shown in its closed configuration. In the closed configuration, a sample 90 containing microorganisms 92 to be analyzed is confined in a thin layer by the two plates. The thickness of the thin layer 90 is adjusted by the two plates and the spacer 40. In this exemplary embodiment, the thin layer thickness 910 is equal to the spacer height, which is selected so that the microorganisms 92 are confined in a single layer between the plates without being compressed. Thus, depending on the suspected effect of the test agent on the microorganism to be analyzed, the microorganisms 92 can be easily observed and monitored by one or more methods. Below, we discuss some exemplary aspects of monitoring microorganisms under the influence of a test agent.

[0087] Figure 3 An exemplary embodiment of the present invention is shown, in which monitoring microorganisms proliferate between two plates. Figures (A) and (B) respectively show the device in a closed configuration at two different time points after a sample containing microorganisms is loaded into the device. The top view of the device is shown on the left, and the cross-sectional view taken along line bb' or cc' is shown on the right. As shown, the QMAX device comprises a first plate 10, a second plate 20, and a spacer 40. Figure (A) shows the device and microorganisms shortly after the two plates enter the closed configuration, wherein the sample is confined in a thin layer by the two plates (the liquid phase component of the sample is not shown), and in this exemplary embodiment, the height of the spacer 40 is selected so that the microorganisms 92 are confined in a monolayer by the two plates. Figure (B) shows a device with microorganisms within a certain period of time after the two plates enter the closed configuration when some microorganisms have begun to split and produce new single microorganisms. For example, the microorganisms 92 represented by the dotted circle in Figure (A) and (B) produce new individuals 94 in Figure (B). It should be noted that although depicted as stationary over time in the figures, microorganisms, particularly living individuals, between the two plates may move on their own despite the fact that in the thin layer the fluid sample is stagnant relative to the plates in the closed configuration.

[0088] Figure 4 Another exemplary embodiment of the present invention is shown, wherein the test agent affects the survival of microorganisms. A top view of the device is shown on the left, and a cross-sectional view taken along line bb', dd' or ee' is shown on the right. Figure (A) depicts a device with Figure 3Figure (A) shows the same situation as Figure (B), while Figures (B) and (C) describe the possible situation that some microorganisms die for a certain period of time after the two plates enter the closed structure. In one case, the individual microorganism indicated by the dotted circle in Figure (A) dies, and its cell integrity is destroyed and appears to "disappear", as represented by the empty dotted circle in Figure (B). In another case, the individual microorganism shown in the dotted square in Figure (A) dies, but it maintains its cell morphology. In this case, the cell viability dye that stains the cell corpses indicates the dead individual 96 in the dotted square in Figure (C).

[0089] In some embodiments, Figure 3 In some embodiments, it is often desirable to monitor cell number, wherein changes in whether and how microorganisms proliferate (increase in number) and / or die-off (decrease in number) are observed under the influence of a test agent, such as, for example, Figure 3 and Figure 4 In some embodiments, it is desirable to monitor cell activity, wherein it is suspected that under the influence of a test agent, it is observed whether and how the microorganisms die but maintain changes in their cell morphology, such as Figure 4 Group (C) shown.

[0090] However, in some embodiments, it is desirable to monitor cell morphology, where it is suspected that their morphology is one aspect affected by the test reagent. In some embodiments, it is desirable to monitor cell motility, where it is suspected that cell motility is one aspect affected by the test reagent. In some embodiments, it is desirable to monitor some other aspect of a microorganism as needed to meet other needs.

[0091] In some embodiments, it may be desirable to monitor any combination of the foregoing aspects of a microorganism.

[0092] An important aspect of the present invention is that the QMAX device is capable of implementing a different range of monitoring possibilities to meet different needs, as will be described in further detail below.

[0093] Reagents on the plate

[0094] To examine the effects of drugs on microorganisms, it is often necessary to contact the microorganisms with various reagents, including test reagents. In some embodiments, the various reagents are added to the sample containing the microorganisms before the sample is deposited on one or both plates. However, one aspect of the present invention provides QMAX devices and methods in which the reagents are coated on one or both plates and added to the sample upon contact with the sample.

[0095] In some embodiments, the sample to be analyzed by the QMAX device is preloaded with a test reagent, and the QMAX device is used solely to monitor microorganisms over time. In some embodiments, the test reagent is coated on the sample contact area of ​​one or both plates and is configured to dissolve into and diffuse into the sample upon contact. In some embodiments, the test reagent comprises at least two parts, one of which is preloaded into the sample and the other of which is coated on the plate.

[0096] In some embodiments, multiplexed testing is performed using a single QMAX device. In some embodiments, different types of test reagents are applied to different locations in the sample contact area of ​​the plate. In some embodiments, the same type of test reagent but at different concentrations is applied to different locations in the sample contact area of ​​the plate. In some embodiments, different test reagents and / or different concentrations of the same test reagent are applied to the plate with fluid separation, for example, a wall structure is present on the plate sample contact area to separate different sub-areas. In some embodiments, there is no fluid separation between the different sub-areas, and the sub-areas are "spaced" from each other due to limited lateral diffusion in the thin layer.

[0097] In certain embodiments, one or two on the plate comprise control area and experimental area on corresponding sample contact area, wherein experimental area comprises one or more test reagents, and test reagent dissolves and is diffused in the sample when contacting sample, and wherein the control area does not comprise this type of test reagent.In certain embodiments, control area and experimental area are fluid isolation.In certain embodiments, control area and experimental area are not fluid isolation, and microorganism can not move through different districts in the time period of inspection.The microorganism in experimental area and control area is compared and is conducive to measuring the impact of test reagent on microorganism.

[0098] In some embodiments, the test agent comprises an antibiotic, such as, but not limited to, actinomycin D, actinomycin, acupuncture, acyclovir, adenine 9-β-D-arabinofuranoside, alamethicone, alamethicone, L-alanine-L-1-aminoethylphosphonic acid, albendazole, 17-(allylamino)-17-demethoxygeldanamycin, astatin, amikacin, amikacin, 7-aminoactinomycin D, 7-aminocephalosporanic acid, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide, (+)-6-aminopenicillanic acid, amoxicillin, amphotericin B, ampicillin, anhydroerythromycin A, anisomycin, antimycin A, antimycin A2, antipain, aphidicolin, aphidicolin, apoptolidin, A, Apramycin, Artesunate, Ascochlorin, Ascomycin, 5-Azacytidine, Azaserine, Azithromycin, Azlocillin, Bacitracin, Antimicrobial Peptides, Bafilomycin A1, Bafilomycin B1, Benstatin, βD-4 Human Recombinant, βD-1 (36 amino acids) Human Recombinant, βD-1 (47 amino acids) Human Recombinant, Bithionol VETRANAL TM , Blasticidin S, Bleomycin, BM-Cyclin, Borrelidin, Brefeldin A, Cyanocyanin A, Calcium ionophore III selector TM , calcium ionophore A23187, calcium ionophore A23187, calcium ionophore A23187, (S)-(+)-camptothecin, capreomycin, carbadox, carbenicillin, carboplatin, cecropin A, cecropin B, cecropin P1, cefaclor, cephalexin VETRANAL TM, cefazolin, cefixime, cefmetazole, cefoperazone, cefotaxime, cefsulodin, ceftazidime, ceftriaxone, cephalexin, cephalosporin, cefotaxime, cephradine, cerosporin, cerulenin, cetylpyridine, chloramphenicol, chlorhexidine, chloroquine, chlortetracycline, chromomycin A3, chrysomexin A, chrysomexin B, cinnamycin, cinoxacin, ciprofloxacin, clarithromycin, clindamycin, clofazimine, clotrimazole, o-cloxacillin, colistin, polyacetylneuraminic acid, compound 48 / 80, concanavalin A, cordycepin, kumarin A1, cryptotanshinone, crystal violet, cyclo Hexamidine, D-cycloserine, cyclosporine A, cyclosporine C, cytochalasin D, cytochalasin B, dacarbazine (antitumor purine analog), daunorubicin, 10-deacetylbaccatin III, dequatacin, defensin HNP-1, defensin HNP-2, demeclocycline hydrochloride, 1-deoxymannonojirimycin, 1-deoxynojirimycin, cis-diammineplatinum(II), 6,9-diamino-2-ethoxyacridine-DL-lactate, cis-diammineplatinum(II), 6-diazo-5-oxo-L-norleucine, 5,7-dichloro-8-hydroxy-2-methylquinoline, dichlorophene Dicloxacillin, diethylcarbamazine, diclofenac, dihydrostreptomycin, dihydrostreptomycin, diloxanide furoate, dimethoprim, dirithromycin, doxorubicin, doxycycline, duramycin, econazole, elastin, anthracene, emetine, enrofloxacin, erythromycin, ethambutol, etoposide, fengyuansu, serine, florfenicol, flubendazole VETRANAL TM, Fluconazole, Flumequine, Flumethasone, 5-fluorocytidine nucleoside analogs, Flurbiprofen cyclooxygenase inhibitors, Metamycin A, Fumagillin, Fumagillin C, Furazolidone, Fusaric acid, G 418, Ganciclovir, Gatifloxacin, Geldamicin, Gentamycin, Gentamycin, Crystal violet, Gliotoxin, Gramicidins, Griseofulvin, Herbimycin A, Cetylpyridine, Magnolia officinalis, Hydrocortisone 21,8-Hydroxyquinoline, 4-Hydroxytamoxifen, (Z)-4-Hydroxytamoxifen, Hygromycin B, Cantharicin, Imipenem, Indomethacin, Indomethacin, Ionomycin, Ir gasan, itraconazole, iturin A, ivermectin, josamycin, K-252a, K-252b, kanamycin, kasugamycin, kendomycin, ketoconazole, flavomycin, L-(+)-lactic acid, lactoferrin B, leptomycin A, leptomycin B, levamisole, levofloxacin, lincomycin, Listeria monocytogenes selective supplement I, LL-37, lomefloxacin, lysostaphin, bombesin I, mebendazole, meclocycline, menadione, 2-thiopyridine N-oxide, N-methyl-1-deoxynojirimycin, 2-methyl-4-isothiazoline-3-hydrochloride, metronidazole, mevastatin, (±)-miconazole, minocycline, mithramycin A, mitomycin C, monensin, morantel, hydroxycarboxyhexanoate, mupirocin, subtilisin, myxothiazole, propylthioamide, nafcillin, naftifine, nalidixic acid, methyl salt Mycin, neocarzinostatin, neomycin, netilmicin, neterotene, niclosamide, nigericin, nikkomycin Z, nisin, nitrofurantoin, nogamycin, no active mycin, norfloxacin, norosin, novobiocin, NP-1, nystatin, ochratoxin A, ofloxacin, oligomycin, oligomycin A, neocyanide, octacycline, oxazolidinone, oxytetracycline, oxytetracycline, paclitaxel, paromomycin, patulin, PD 404,182, pediocin, pefloxacin, D-penicillamine, penicillin G, penicillin V, pentamidine, PGLa, 1,10-Phenanthroline, phenazine, penicillin V, phleomycin, fosfomycin, pimaricin, pipemidic acid, piperacillin, pirarubicin, plate mycin, polymyxin B, poly(vinylpyrrolidone) iodine complex, potassium clavulanate:cellulose (1:1), potassium sorbate, praziquantel, puromycin, pyrantel, pyrazinamide, bisquinacridone, pyrrolocin, quinine, 8-quinolinol, radicicol, ramoplanin, rapamycin, phaeomycin, rafamycin A, ribavirin, ribostamycin, albendazole sulfoxide, rifabutin, rifampicin, rifamycin SV, rifapentine, rifaximin, ristocetin, rolicycline, roxithromycin , Salinomycin, Sanamycin, Sinafenac, Perillomycin, Sorbic acid, Sordolin, Sparfloxacin, Spectinomycin, Spergualin, Spiramycin, Staurosporine, Streptolysin O, Streptomycin, Streptomycin, Streptomycin melanin, Streptozotocin, Succinylsulfathiazole, Sulconazole, Sulfadimyl, Sulfadiazine, Sulfadiazine, Sulfadiazine, Sulfadimethoxine, Sulfadiaz ... E, tamoxifen, tazobactam, teicoplanin, terbinafine, terconazole, tetracycline, tetramidazole, thiabendazole, thiamphenicol, sodium thimerosal, thioglucosidone, thiostrepton, thio-TEPA, thymol, tiamulin, hydroxythiazolin, tioconazole, tobramycin, aminoglycoside antibiotics, tobramycin, tolnaftate, toyocamycin, triazocine C, trichlorfon, trimethoprim, tuberculin, tunicamycin, tunicamycin C2 homologue, tylosin, valacyclovir, valinomycin, vinblastine, virginiamycin S1, virginiamycin M1, and any analogs, salts and derivatives thereof.

[0099] In some embodiments, the test agent includes an antifungal agent such as, but not limited to, polyene antifungals (amphotericin B, candidiasis, filipin, harstomycin, natamycin, nystatin, chelamycin), imidazoles (e.g., bifonazole, butoconazole, clotrimazole, econazole, fenticonazole, isoconazole, ketoconazole, luliconazole, miconazole, omoconazole, oxiconazole, sertaconazole, sulconazole, tioconazole), triazoles (albaconazole, epoxiconazole, fluconazole, Isavuconazole, itraconazole, posaconazole, propiconazole, ravuconazole, terconazole, voriconazole), thiazoles (e.g., abafungin), allylamines (e.g., amorolfine, butenafine, naftifon and terbinafine), echinocandins (e.g., anidulafungin, caspofungin and micafungin), aurone, benzoic acid, ciclopirox, flucytosine and 5-fluorocytosine, griseofulvin, chloropropynyl iodide, tolnaftate, undecylenic acid, crystal violet, Peruvian balsam, analogs thereof, salts thereof and derivatives thereof.

[0100] In addition to or in place of test reagents, in some embodiments, the QMAX device includes on one or both plates various other reagents to be added to the sample, such as, but not limited to, staining dyes to label microorganisms (cell viability dyes as well as many others), life support reagents to promote microbial survival (e.g., nutrients, oxygen sources, pH buffers), and proliferation reagents to maintain or promote microbial proliferation (e.g., hormones, siderophores).

[0101] In some embodiments, cell viability dyes include, but are not limited to, propidium iodide (PI), 7-AAD (7-aminoactinomycin D), trypan blue, calcein violet AM, calcein AM, fixable viability dyes (FVD) conjugated to different fluorophores, SYTO9 and other nucleic acid dyes, resazurin and colored formazan (MTT / XTT) and other mitochondrial dyes, and any combination thereof, etc. In some embodiments, the sample contains bacteria and it is necessary to determine the viability of the bacteria in the sample, and the device further includes a bacterial viability dye, such as PI, SYTO9, etc., on one or both plates to differentially stain live cells from dead cells.

[0102] In some embodiments, the dye is selected from the group consisting of acid fuchsin, alcian blue 8GX, alizarin red S, aniline blue WS, auramine O, azocarmine B, azocarmine G, sky blue A, sky blue B, sky blue C, basic fuchsin, Bismarck brown Y, brilliant cresyl blue, brilliant green, carmine, chlorazole black E, Congo red, CI cresyl violet, crystal violet, Darrow red, eosin B, eosin Y, erythrosine, ethyl eosin, ethyl green, fast green FCF, fluorescein isothiocyanate, Giemsa dye, hematoxylin, hematoxylin & eosin, indigo carmine, Janus Green B, MacNeal's dye 1899, Light Green SF, Malachite Green, Matthew's Yellow, Methyl Orange, Methyl Violet 2B, Methylene Blue, Methylene Blue, Methylene Violet, (Methylene Blue), Neutral Red, Aniline Black, Nile Blue A, Nuclear Fast Red, Oil Red, Orange G, Orange II, Orcein, Parafuchsin, Bamboo Red B, Strong Protein Silver S, Pyronine B, Pyronine, Resazurin, Rose Bengal, Safranin O, Sudan Black B, Sudan III, Sudan IV, Tetrachrome Stain (MacNeal), Lowe's Violet, Toluidine Blue, Weigert, Wright's Stain, and any combination thereof.

[0103] In some embodiments, the dye is conjugated to a fluorescent molecule (fluorophore), including but not limited to, IRDye800CW, Alexa790, Dylight 800, fluorescein, fluorescein isothiocyanate, succinimidyl ester of carboxyfluorescein, succinimidyl ester of fluorescein, 5-isomer of fluorescein dichlorotriazine, caged carboxyfluorescein-alanine-carboxamide, Oregon Green 488, Oregon Green 514; fluorescein yellow, acridine orange, rhodamine, tetramethylrhodamine, Texas Red, propidium iodide, JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide), tetrabromorhodamine 123, rhodamine 6G, TMRM (tetramethylrhodamine methyl ester), TMRE (tetramethylrhodamine ethyl ester), tetramethylrhodamine, rhodamine B and 4-dimethylaminotetramethylrhodamine, Green Color fluorescent protein, blue-shifted green fluorescent protein, blue-green-shifted green fluorescent protein, red-shifted green fluorescent protein, yellow-shifted green fluorescent protein, 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid; acridine and derivatives, e.g., acridine, acridine isothiocyanate; 5-(2'-aminoethyl)aminonaphthalene-1-sulfonic acid (EDANS); 4-amino-N-[3-vinylsulfonyl)phenyl]naphthamide-3,5-disulfonate; N-(4-anilino-1-naphthyl)maleimide; anthranilamide; 4,4-difluoro-5-(2-thienyl)-4-boron-3a,4a-diaza-5-indacene-3-propionic acid BODIPY; Cascade Blue; Brilliant Yellow; Coumarins and derivatives: Coumarin, 7-amino-4-methylcoumarin (AMC, Coumarin 120), 7-amino-4-methylcoumarin (Coumarin 151); cyanine dyes; cyanosine; 4',6-diamino-2-phenylindole (DAPI); 5',5"-dibromopyrogallolsulfonaphthalene (bromopyrogallol red); 7-diethylamino-3-(4'-isothiocyanatophenyl)-4-methylcoumarin; diethylenetriamine pentaacetate; 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid; 4,4'-diisothiocyanato-2,2'-yldisulfonic acid disodium salt; 5-(dimethylamino)naphthalene-1-sulfonyl chloride (DNS, dansyl) Chloro-4'-dimethylaminophenylazophenyl-4'-isothiocyanate (DABITC); Eosin and derivatives: Eosin, Eosin isothiocyanate, Erythrosine and derivatives: Erythrosine B, Erythrosine, isothiocyanate; Ethidium; Fluorescein and its derivatives: 5-carboxyfluorescein (FAM), 5-(4,6-dichlorotriazin-2-yl)amino-fluorescein (DTAF), 2',7'-dimethoxy-4',5'-dichloro-6-carboxyfluorescein (JOE), Fluorescein, Fluorescein isothiocyanate, QFITC, (XRITC); Fluorescein amine; IR144; IR1446; Malachite green isothiocyanate; 4-Methylumbelliferyl o-cresolphthalein; Nitrotyrosine; Parafuchsin; Phenol red;B-phycoerythrin; o-phthalaldehyde; pyrene and its derivatives: pyrene, pyrenebutyric acid, succinimidyl 1-pyrene; butyrate quantum dots; Reactive Red 4 (Cibacron; TM Brilliant Red 3B-A) Rhodamine and derivatives: 6-carboxy-X-rhodamine (ROX), 6-carboxyrhodamine (R6G), lissamine rhodamine B sulfonyl chloride rhodamine (Rhod), Rhodamine B, Rhodamine 123, Rhodamine X isothiocyanate, sulforhodamine B, sulforhodamine 101, sulfonyl chloride derivatives of 5-sulforhodamine (Texas Red); N,N,N',N'-tetramethyl-6-carboxyrhodamine TAMRA; tetramethylrhodamine; tetramethylrhodamine isothiocyanate (TRITC); riboflavin; 5-(2'-aminoethyl)aminonaphthalene-1-sulfonic acid (EDANS), 4-(4'-dimethylaminophenylazo)benzoic acid (DABCYL), rosocyanic acid; CAL Fluorescent Orange 560; terbium chelate derivatives; Cy3; Cy5; Cy5.5; Cy7; IRD700; IRD 800; La Jolla Blue; phthalocyanines; and naphthalocyanines, coumarins and related dyes, xanthene dyes, such as rhodols, resorufins, bimanes, acridines, isoindoles, dansyl dyes, aminophthalhydrazides (e.g., luminol), and isoluminol derivatives, aminophthalimides, aminonaphthalimides, aminobenzofurans, aminoquinolines, dicyanohydroquinones, fluorescent europium and terbium complexes; combinations thereof, and the like. Suitable fluorescent and chromogenic proteins include, but are not limited to, green fluorescent protein (GFP), including, but not limited to, GFP from Aequoria victoria or a derivative thereof, e.g., a "humanized" derivative, such as enhanced GFP; GFP from another species, e.g., Renilla reniformis, Renilla mulleri, or Ptilosarcus guernyi; "humanized" recombinant GFP (hrGFP); any of a variety of fluorescent and colored proteins from coral species; any combination thereof; and the like.

[0104] Imaging and image processing

[0105] In some embodiments, examination of the effect of a drug on a microorganism is accomplished by monitoring the microorganisms in a uniform thickness layer and determining the effect of the test agent on the microorganisms when the two plates are in a closed configuration as described above.

[0106] In some embodiments, monitoring is accomplished optically, and one or both of the plates are transparent, enabling optical inspection and / or imaging of microorganisms between the two plates.

[0107] In some embodiments, microscopy is used to monitor microorganisms in the device. In some embodiments, a standard laboratory microscope is used, while in some embodiments, a miniaturized or custom-designed microscope is used.

[0108] In some embodiments, monitoring is achieved by imaging. An imager is used to obtain images of signals emitted by microorganisms in the device. In some embodiments, the imager comprises a camera and a light source for imaging the microorganisms in the thin layer.

[0109] In some embodiments, a series of images are taken of the microorganisms in the device at discrete time points. In some embodiments, images of the microorganisms in the device are taken over a period of time.

[0110] In some embodiments, visual inspection of the microorganisms is performed by a professional or non-professional user of the device with the aid of a microscope, images obtained of the microorganisms, and / or videos obtained.

[0111] In some embodiments, a processor is configured to process the acquired images of microorganisms and / or the acquired videos. The processor includes electronics, a signal processor, and hardware and software for receiving and processing images and identifying and analyzing platelets in the images.

[0112] In some embodiments, the monitoring and determining steps comprise:

[0113] i. obtaining a series of images of a plate containing microorganisms in a sample layer of uniform thickness between the plates at discrete time points;

[0114] ii. identifying and / or counting microorganisms in at least one region of each image; and

[0115] iii. Assessing changes in microorganisms between at least some discrete time points.

[0116] In some embodiments, the monitoring and determining steps comprise:

[0117] i. a video comprising a series of images of a plate containing microorganisms in a layer of uniform thickness, obtained at discrete time points during a first time period;

[0118] ii. identifying and tracking at least a portion of the microorganisms in the video; and

[0119] iii. Evaluating changes in the tracked microorganisms during at least a portion of the first time period.

[0120] In some embodiments, the identifying step comprises identifying microorganisms stained with a dye, and wherein the dye comprises a viability dye that indicates whether the stained cells are alive or dead.

[0121] In some embodiments, the change in microorganisms includes, but is not limited to, a change in the total number of microorganism individuals, a change in microorganism concentration, a change in microorganism morphology, a change in microorganism motility, a change in microorganism absorption of staining dyes, and any combination thereof.

[0122] In some embodiments, changes in the number or concentration of individual microorganisms are evaluated. The processor is configured to count the number of identified microorganisms in at least one region of each image. In some cases, the concentration of the microorganisms is determined. An advantage of the present invention is that, in some embodiments, the spacers control the thickness of the final sample layer to be very uniform. Therefore, by multiplying the region of interest by the final sample thickness, the volume of the sample in at least one region of each image can be easily calculated, and the concentration of the microorganisms in the region of interest can be easily derived.

[0123] In some embodiments, changes in the number of individual microorganisms are assessed by tracking at least a portion of the microorganisms in the acquired video. When tracking individual microorganisms, the processor is further configured to identify events when the tracked individuals undergo cell division (producing new individuals and increasing their number) and / or disappear due to cell death and cell body decomposition.

[0124] In some embodiments, changes in microbial morphology are assessed by analyzing geometric parameters of images of each individual microorganism in the acquired images and / or videos.

[0125] In some embodiments, changes in microbial motility are assessed by analyzing movement parameters of tracked microbial individuals in acquired videos, such as, but not limited to, movement speed, direction, and duration and frequency of immobility.

[0126] In some embodiments, changes in the uptake of a staining dye by a microorganism are assessed by evaluating the signal emitted by the staining dye within the boundaries of individual microorganisms.

[0127] System for examining the effects of drugs on microorganisms

[0128] Another aspect of the present invention is to provide a system for examining the effects of drugs on microorganisms. In certain embodiments, the system allows for remote health monitoring, consultation, and the like.

[0129] In some embodiments, the system includes:

[0130] (a) QMAX device;

[0131] (b) an imager comprising a camera and a light source for imaging microorganisms in a layer of uniform thickness; and

[0132] (c) A processor comprising electronics, a signal processor, and hardware and software for receiving and processing images and identifying and analyzing microorganisms in the images.

[0133] In some embodiments, the imager is configured to obtain a series of images of the microorganism at discrete points in time.

[0134] In some embodiments, the processor is configured to receive and analyze the series of images, identifying and counting microorganisms in a first region of each image.

[0135] In some embodiments, the imager is configured to obtain a video comprising a series of images of the microorganisms over a specific time period.

[0136] In some embodiments, the processor is configured to receive and analyze the video, identify and track at least a portion of the microorganisms in the video, and evaluate changes in the tracked microorganisms over at least a portion of the first time period.

[0137] In some embodiments, the mobile communication device is used as an imager and optionally as an image processor. In some embodiments, the system includes:

[0138] (a) A QMAX device as described in any preceding or following embodiment;

[0139] (b) A mobile communication device comprising:

[0140] i. an imager comprising one or more cameras for imaging microorganisms in a sample;

[0141] ii. a processor comprising electronics, a signal processor, and hardware and software for receiving and / or processing images of microorganisms and for remote communication; and

[0142] (c) a light source from the mobile communication device or an external source, wherein the light source is configured to provide illumination to the sample for imaging with the camera.

[0143] In some embodiments, the system further comprises:

[0144] (d) A housing configured to hold a sample and to be mounted to the mobile communication device.

[0145] In some embodiments, the housing includes optics for facilitating imaging and / or signal processing of a sample by a mobile communication device, and a mount configured to hold the optics on the mobile communication device.

[0146] In some embodiments, the mobile communication device is configured to transmit the test results to medical personnel, a medical institution, or an insurance company.

[0147] In some embodiments, the mobile communication device is further configured to communicate information about the subject with medical personnel, medical institutions, or insurance companies. In some embodiments, the mobile communication device is configured to receive a prescription, diagnosis, or recommendation from a medical professional. In some embodiments, the mobile communication device communicates with a remote location via WiFi or a cellular network.

[0148] In some embodiments, the mobile communication device is a mobile phone.

[0149] application

[0150] The present invention has particular use in many aspects of a clinical / diagnostic setting.

[0151] In some embodiments, the present invention can be used for susceptibility testing, wherein the effects of antimicrobial drugs on potential microbial pathogens causing patient infections are screened to help medical personnel select the most effective antimicrobial drug or treatment strategy to treat the patient's infection. In some embodiments, testing is performed to facilitate identification of pathogenic microorganisms. In some embodiments, the QMAX device, system, and method are used to examine the effects of antimicrobial drugs on potential microorganisms causing conditions in patients or other subjects. In some embodiments, high-throughput testing of various antimicrobial drugs at various concentrations / doses against potential microbial pathogens is performed in parallel using a single device or system configured with multiplexing functionality as described above.

[0152] Table 1 is a chart of common pathogens. In some embodiments, the present invention can be used to test the effects of a drug on one or more of the bacteria listed in the chart.

[0153] Table 1 Common pathogens

[0154]

[0155]

[0156]

[0157] In certain embodiments, the present invention can be used for many other tests related to biological and / or clinical samples except sensitivity testing.These samples include but are not limited to cells, tissues and body fluids.The body fluid of interest includes but is not limited to amniotic fluid, aqueous humor, vitreous humor, blood (for example, whole blood, fractionated blood, plasma, serum), breast milk, cerebrospinal fluid (CSF), cerumen (ear wax), chyle, chyme, endolymph, perilymph, excreta, gastric acid, gastric juice, lymph, mucus (including nasal drainage and sputum), pericardial fluid, peritoneal fluid, pleural fluid, pus, rheumatic fluid, saliva, sebum (skin oil), semen, sputum, sweat, synovial fluid, tears, vomitus, urine and exhaled condensate.In certain embodiments, the sample comprises human body fluid. In some embodiments, the sample comprises at least one of: cells, tissue, body fluid, feces, amniotic fluid, aqueous humor, vitreous humor, blood, whole blood, fractionated blood, plasma, serum, breast milk, cerebrospinal fluid, cerumen, chyle, chyme, endolymph, perilymph, feces, gastric acid, gastric juice, lymph, mucus, nasal drainage, sputum, pericardial fluid, peritoneal fluid, pleural fluid, pus, rheumatic fluid, saliva, sebum, semen, sputum, sweat, synovial fluid, tears, vomitus, urine, and exhaled condensate.

[0158] In some embodiments, the present invention can be used for many other tests involving microorganisms, such as, but not limited to, environmental testing, food testing, and forensic testing. Environmental samples can be obtained from any suitable source, such as rivers, oceans, lakes, rainwater, snow, sewage, sewage treatment runoff, agricultural runoff, industrial runoff, tap water or drinking water, etc. Food samples can be obtained from any suitable source, such as tap water, drinking water, prepared foods, processed foods, or unprocessed foods, etc.

[0159] In some embodiments, the sample is obtained directly from a source. In some embodiments, the sample is pre-loaded with reagents or pre-treated as needed before being deposited on the plate of the device. In some embodiments, when it is desired to increase the level of the microorganism being analyzed, the sample to be deposited on the device comprises a product from a sample culture obtained directly from the source.

[0160] Control plate spacing and sample thickness using spacers

[0161] According to the invention, the spacing between the two plates and thus the sample thickness is controlled by using spacers.

[0162] Spacer height. In some embodiments, all spacers have the same predetermined height. In some embodiments, the spacers have different predetermined heights. In some embodiments, the spacers can be divided into groups or regions, where each group or region has its own spacer height. And in some embodiments, the predetermined height of the spacers is the average height of the spacers. In some embodiments, the heights of the spacers are approximately the same. In some embodiments, a percentage of the spacers have the same height.

[0163] The height of the spacer is selected by the desired adjustment spacing between the plates and / or the final sample thickness and the remaining sample thickness of the adjustment. The spacer height (predetermined spacer height), the spacing between the plates and / or the sample thickness is 3 nm or less, 10 nm or less, 50 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 800 nm or less, 1000 nm or less, 1 μm or less, 2 μm or less, 3 μm or less, 5 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, 500 μm or less, 800 μm or less, 1 mm or less, 2 mm or less, 4 mm or less, or in a range between any two of these values.

[0164] The spacer height, spacing between plates and / or sample thickness is from 1 nm to 100 nm in one preferred embodiment, from 100 nm to 500 nm in another preferred embodiment, from 500 nm to 1000 nm in a separate preferred embodiment, from 1 μm (i.e. 1000 nm) to 2 μm in another preferred embodiment, from 2 μm to 3 μm in a separate preferred embodiment, from 3 μm to 5 μm in another preferred embodiment, from 5 μm to 10 μm in a separate preferred embodiment, and from 10 μm to 50 μm in another preferred embodiment, and from 50 μm to 100 μm in a separate preferred embodiment.

[0165] In some embodiments, the spacer height is precisely controlled. The relative accuracy of the spacers (i.e., the ratio of the deviation to the desired spacer height) is 0.001% or less, 0.01% or less, 0.1% or less, 0.5% or less, 1% or less, 2% or less, 5% or less, 8% or less, 10% or less, 15% or less, 20% or less, 30% or less, 40% or less, 50% or less, 60% or less, 70% or less, 80% or less, 90% or less, 99.9% or less, or a range between any two of these values.

[0166] In some embodiments, the spacer height, spacing between plates, and / or sample thickness is: (i) equal to or slightly larger than the smallest dimension of the analyte, or (ii) equal to or slightly larger than the largest dimension of the analyte. "Slightly larger" means about 1% to 5% larger, and any value in between.

[0167] In some embodiments, the spacer height, spacing between plates, and / or sample thickness is greater than the smallest dimension of the analyte (eg, the analyte has an anisotropic shape), but less than the largest dimension of the analyte.

[0168] For example, a red blood cell has a disc shape with a minimum dimension of 2 μm (disc thickness) and a maximum dimension of 11 μm (disc diameter). In an embodiment of the present invention, the spacers are selected so that the spacing between the inner surfaces of the plates in the relevant area is 2 μm (equal to the minimum dimension) in one embodiment, 2.2 μm in another embodiment, or 3 μm (50% larger than the minimum dimension) in another embodiment, but less than the maximum dimension of the red blood cell. Such an embodiment has certain advantages in blood cell counting. In one embodiment, for red blood cell counting, by having the inner surface spacing be 2 μm or 3 μm and any value in between, an undiluted whole blood sample is confined to this spacing; on average, each red blood cell (RBC) does not overlap with another, allowing for accurate visual counting of the red blood cells. (Excessive overlap between RBCs can lead to significant errors in the count).

[0169] In some embodiments, the spacer height, spacing between plates, and / or sample thickness is: (i) equal to or slightly smaller than the smallest dimension of the analyte, or (ii) equal to or slightly smaller than the largest dimension of the analyte. "Slightly smaller" means that it is about 1% to 5% smaller, and any number in between.

[0170] In some embodiments, the spacer height, spacing between plates, and / or sample thickness is greater than the smallest dimension of the analyte (eg, the analyte has an anisotropic shape), but less than the largest dimension of the analyte.

[0171] In the present invention, in some embodiments, the plates and spacers are used not only to adjust the thickness of the sample, but also to adjust the orientation and / or surface density of the analytes / entities in the sample when the plate is in a closed configuration. When the plate is in a closed configuration, a thinner thickness of the sample results in fewer analytes / entities per surface area (i.e., a smaller surface concentration).

[0172] Spacer Transverse Dimensions. For open spacers, the transverse dimensions can be characterized by their transverse dimensions in two orthogonal directions, x and y (sometimes referred to as width). The transverse dimensions of the spacer in each direction can be the same or different. In some embodiments, the lateral dimension in each direction (x or y) is 1 nm or less, 3 nm or less, 5 nm or less, 7 nm or less, 10 nm or less, 20 nm or less, 30 nm or less, 40 nm or less, 50 nm or less, 100 nm or less, 200 nm or less, 500 nm or less, 800 nm or less, 1000 nm or less, 1 μm or less, 2 μm or less, 3 μm or less, 5 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 50 μm or less, 100 μm or less, 150 μm or less, 200 μm or less, 300 μm or less, or 500 μm or less, or in a range between any two of these values.

[0173] In some embodiments, the ratio of the lateral dimensions in the x and y directions is 1, 1.5, 2, 5, 10, 100, 500, 1000, 10,000, or a range between any two of the values. In some embodiments, different ratios are used to adjust the direction of sample flow; the larger the ratio, the flow is along one direction (the direction of the larger dimension).

[0174] In some embodiments, the different lateral dimensions of the spacers in the x and y directions serve to (a) use the spacers as scale markers to indicate plate orientation, (b) use the spacers to generate more sample flow in a preferred direction, or both.

[0175] In a preferred embodiment, the period, width and height of the spacers are substantially the same. In some embodiments, all spacers have the same shape and size. In some embodiments, the spacers have different lateral dimensions.

[0176] For the enclosing spacer, in some embodiments, the inner lateral shape and dimensions are selected based on the total volume of the sample to be enclosed by the enclosing spacer, where the volume dimensions have been described in the present disclosure; and in certain embodiments, the outer shape and dimensions are selected based on the required strength to support the pressure of the liquid against the spacer and the compressive pressure of the pressing plate.

[0177] In certain embodiments, the aspect ratio of the height to the average lateral dimension of the column spacers is 100,000, 10,000, 1,000, 100, 10, 1, 0.1, 0.01, 0.001, 0.0001, 0,00001, or a range between any two of these values.

[0178] Spacer spacing. The spacer can be a single spacer or a plurality of spacers on the plate or in the sample-related area. In some embodiments, the spacers on the plate are configured and / or arranged in an array, and the array is fixed-spaced, a non-fixed-spaced array, or fixed-spaced at some locations on the plate and non-fixed-spaced at other locations.

[0179] In some embodiments, the spaced array of spacers is arranged in a lattice of squares, rectangles, triangles, hexagons, polygons, or any combination thereof, where combination means that different locations of the plate have different spacer lattices.

[0180] In some embodiments, the spacer spacing of the spacer array is fixedly spaced in at least one direction of the array (ie, uniform spacer spacing). In some embodiments, the spacer spacing is configured to improve uniformity between plate spacing in the closed configuration.

[0181] In some embodiments, the distance between adjacent spacers (i.e., the spacer pitch) is 1 μm or less, 5 μm or less, 7 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 40 μm or less, 50 μm or less, 60 μm or less, 70 μm or less, 80 μm or less, 90 μm or less, 100 μm or less, 200 μm or less, 300 μm or less, 400 μm or less, or in a range between any two of said values.

[0182] In certain embodiments, the spacer spacing is 400 μm or less, 500 μm or less, 1 mm or less, 2 mm or less, 3 mm or less, 5 mm or less, 7 mm or less, 10 mm or less, or any range therebetween. In certain embodiments, the spacer spacing is 10 mm or less, 20 mm or less, 30 mm or less, 50 mm or less, 70 mm or less, 100 mm or less, or any range therebetween.

[0183] The distance between adjacent spacers (i.e., the spacer spacing) is selected so that for given properties of the plate and sample, in the closed configuration of the plate, in some embodiments, the sample thickness between two adjacent spacers varies by at most 0.5%, 1%, 5%, 10%, 20%, 30%, 50%, 80%, or within any range between the stated values; or in certain embodiments, by at most 80%, 100%, 200%, 400%, or within a range between any two of the stated values.

[0184] Obviously, in order to maintain a given sample thickness variation between two adjacent spacers, a closer spacer spacing is required when using a more flexible plate.

[0185] In a preferred embodiment, the spacers are a regularly spaced square array, wherein the spacers are pillars having a height of 2-4 μm, an average lateral dimension of 1-20 μm, and a spacer pitch of 1 μm-100 μm.

[0186] In a preferred embodiment, the spacers are a regularly spaced square array, wherein the spacers are pillars having a height of 2-4 μm, an average lateral dimension of 1-20 μm, and a spacer pitch of 100 μm-250 μm.

[0187] In a preferred embodiment, the spacers are a regularly spaced square array, wherein the spacers are pillars having a height of 4-50 μm, an average lateral dimension of 1-20 μm, and a spacer pitch of 1 μm-100 μm.

[0188] In a preferred embodiment, the spacers are a regularly spaced square array, wherein the spacers are pillars having a height of 4-50 μm, an average lateral dimension of 1-20 μm, and a spacer pitch of 100 μm-250 μm.

[0189] The spacing of the spacer array is 1 nm to 100 nm in one preferred embodiment, 100 nm to 500 nm in another preferred embodiment, 500 nm to 1000 nm in a separate preferred embodiment, 1 μm (i.e., 1000 nm) to 2 μm in another preferred embodiment, 2 μm to 3 μm in a separate preferred embodiment, 3 μm to 5 μm in another preferred embodiment, 5 μm to 10 μm in a separate preferred embodiment, 10 μm to 50 μm in another preferred embodiment, 50 μm to 100 μm in a separate preferred embodiment, 100 μm to 175 μm in a separate preferred embodiment, and 175 μm to 300 μm in a separate preferred embodiment.

[0190] Spacer Density. The spacers are arranged on the respective plates at a surface density of greater than 1 per μm², greater than 1 per 10 μm², greater than 1 per 100 μm², greater than 1 per 500 μm², greater than 1 per 1000 μm², greater than 1 per 5000 μm², greater than 1 per 0.01 mm², greater than 1 per 0.1 mm², greater than 1 per 1 mm², greater than 1 per 5 mm², greater than 1 per 10 mm², greater than 1 per 100 mm², greater than 1 per 1000 mm², greater than 1 per 10,000 mm², or within a range between any two of these values. In some embodiments, the spacers have a density of at least 1 / mm², at least 10 / mm², at least 50 / mm², at least 100 / mm², at least 1,000 / mm², or at least 10,000 / mm².

[0191] The spacer area fill factor is defined as the ratio of the spacer area to the total area or the ratio of the spacer spacing to the width. In some embodiments, the fill factor is at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, or a range between any two of the values. In certain embodiments, the fill factor is at least 2.3%.

[0192] In a device comprising two plates and a spacer, the fourth power of the spacer distance (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD4 / (hE)) is 5×106 μm3 / GPa or less.

[0193] In a device comprising two plates and a spacer, the fourth power of the spacer distance (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD4 / (hE)) is 5×105 μm3 / GPa or less.

[0194] In a device comprising two plates and spacers, the spacers have a columnar shape, a substantially flat top surface, a predetermined substantially uniform height, and a predetermined constant spacer spacing, the spacer spacing being at least about 2 times larger than the size of the analyte, wherein the Young's modulus of the spacer multiplied by the filling factor of the spacer is equal to or greater than 2 MPa, wherein the filling factor is the ratio of the spacer contact area to the total plate area, and wherein for each spacer, the ratio of the lateral dimension of the spacer to its height is at least 1 (one).

[0195] In a device comprising two plates and spacers, the spacers have a columnar shape, a substantially flat top surface, a predetermined substantially uniform height, and a predetermined constant spacer spacing, the spacer spacing being at least about 2 times greater than a size of the analyte, wherein the Young's modulus of the spacer multiplied by the filling factor of the spacer is equal to or greater than 2 MPa, wherein the filling factor is the ratio of the spacer contact area to the total plate area, and wherein for each spacer, the ratio of the lateral dimension of the spacer to its height is at least 1 (one), wherein the fourth power of the spacer spacing (ISD) divided by the thickness (h) and Young's modulus (E) of the flexible plate (ISD4 / (hE)) is 5x106μm3 / GPa or less.

[0196] In a device including two plates and spacers, a ratio of a spacer pitch of the spacers to an average width of the spacers is 2 or greater, and a filling factor of the spacers multiplied by a Young's modulus of the spacers is 2 MPa or greater.

[0197] Examples of the Invention

[0198] A1. An apparatus for examining the effects of a drug on a microorganism, comprising:

[0199] A first plate, a second plate, and a spacer, wherein:

[0200] i. The plates are movable relative to each other into different configurations, including an open configuration and a closed configuration;

[0201] ii. each plate has a sample contact area on its respective sample surface for contacting a sample, wherein the sample contains the microorganisms to be analyzed, and

[0202] iii. one or both of the plates contain spacers fixed to the respective sample contact areas, the spacers having a predetermined height that is uniform, and at least one of the spacers is within the sample contact area,

[0203] wherein in an open configuration, the two plates are partially or completely separated, the spacing between the plates is not adjusted by spacers, and the sample is deposited on one or both of the plates;

[0204] Wherein in the closed configuration, the closed configuration is configured after the sample is deposited in the open configuration: at least a portion of the sample is compressed by two plates into a layer of very uniform thickness, and the uniform thickness of the layer is defined by the sample contacting surfaces of the plates and adjusted by the plates and spacers.

[0205] B0. A system for examining the effects of a drug on a microorganism, comprising:

[0206] (a) The device described in any one of the embodiments;

[0207] (b) an imager comprising a camera and a light source for imaging microorganisms in a layer of uniform thickness; and

[0208] (c) A processor comprising electronics, a signal processor, and hardware and software for receiving and processing images and identifying and analyzing microorganisms in the images.

[0209] B1. A system for examining the effects of a drug on a microorganism, comprising:

[0210] (a) The device described in any one of the embodiments;

[0211] (b) a mobile communication device comprising:

[0212] i. an imager comprising one or more cameras for imaging microorganisms in a sample;

[0213] ii. a processor comprising electronics, a signal processor, and hardware and software for receiving and / or processing images of microorganisms and for remote communication; and

[0214] (c) a light source from the mobile communication device or an external source, wherein the light source is configured to provide illumination to the sample for imaging with the camera.

[0215] C1. A method for examining the effects of a drug on a microorganism, comprising the following steps:

[0216] (a) obtaining a sample containing a microorganism to be analyzed and mixing it with a test reagent;

[0217] (b) obtaining a first plate and a second plate movable relative to each other into different configurations, the different configurations comprising an open configuration and a closed configuration, wherein:

[0218] i. Each plate has a sample contact area on its respective surface for contacting the sample, and

[0219] ii. one or both of the plates contain spacers fixed to the respective sample contact surfaces,

[0220] wherein the spacers have a predetermined substantially uniform height, and at least one of the spacers is within the sample contact area;

[0221] (c) depositing a sample on one or both of the plates while the plates are in the open configuration,

[0222] wherein in the open configuration, the two plates are partially or completely separated and the spacing between the plates is not adjusted by spacers;

[0223] (d) after (c), joining the two plates together and pressing the plates into a closed configuration, wherein in the closed configuration: at least a portion of the sample is compressed by the two plates into a layer of very uniform thickness, the uniform thickness of the layer being defined by the sample surfaces of the two plates and adjusted by the spacers and the plates;

[0224] (e) When the plate is in the closed configuration, monitoring the microorganisms in the uniform thickness layer and determining the effect of the test agent on the microorganisms.

[0225] CC1. A method for examining the effects of a drug on a microorganism, comprising the following steps:

[0226] (a) obtaining a sample containing microorganisms to be analyzed;

[0227] (b) obtaining a first plate and a second plate movable relative to each other into different configurations, the different configurations comprising an open configuration and a closed configuration, wherein:

[0228] i. Each plate has a sample contact area on its respective surface for contacting the sample,

[0229] ii. One or both of the plates contain a spacer fixed to the corresponding sample contact surface, and

[0230] iii. at least one of the plates contains a test reagent on a corresponding sample contact area, the test reagent dissolving and diffusing in the sample upon contact with the sample,

[0231] wherein the spacers have a predetermined substantially uniform height, and at least one of the spacers is within the sample contact area;

[0232] (c) depositing a sample on one or both of the plates while the plates are in the open configuration,

[0233] wherein in the open configuration, the two plates are partially or completely separated and the spacing between the plates is not adjusted by spacers;

[0234] (d) after (c), joining the two plates together and pressing the plates into a closed configuration, wherein in the closed configuration: at least a portion of the sample is compressed by the two plates into a layer of very uniform thickness, the uniform thickness of the layer being defined by the sample surfaces of the two plates and adjusted by the spacers and the plates;

[0235] (e) When the plate is in the closed configuration, monitoring the microorganisms in the uniform thickness layer and determining the effect of the test agent on the microorganisms.

[0236] A2. The device, system, or method of any preceding embodiment, wherein one or both of the plates comprises one or more test reagents on the respective sample contact areas, the test reagents dissolving and diffusing into the sample upon contact with the sample.

[0237] A3. An apparatus, system, or method as described in any preceding embodiment, wherein one or both of the plates comprises a control zone and an experimental zone on the corresponding sample contact area, wherein the experimental zone comprises one or more test reagents that dissolve and diffuse into the sample upon contact with the sample, and wherein the control zone does not comprise such test reagents.

[0238] A4. The device, system, or method of any preceding embodiment, wherein the spacers have a uniform height equal to or less than the average size of the microorganisms.

[0239] A5. The device, system, or method of any preceding embodiment, wherein the uniform height of the spacers is in the range of 75% to 125% of the average size of the microorganisms.

[0240] A6. The apparatus, system, or method of any preceding embodiment, wherein at least one of the plates is transparent.

[0241] A7. The device, system, or method of any preceding embodiment, wherein one or both of the plates comprises a dye on the corresponding sample contact area that dissolves in the sample and stains the microorganisms when in contact with the sample.

[0242] A8. The device, system or method of embodiment A4, wherein the dye is fluorescently labeled.

[0243] A9. An apparatus, system or method as described in any of the preceding embodiments, wherein one or both of the plates comprises a vital dye on the corresponding sample contact area, the vital dye being selected from the group consisting of: propidium iodide, 7-AAD, trypan blue, calcein violet AM, calcein AM, fixable vital dyes, SYTO9 and other nucleic acid dyes, resazurin and colored formazan (MTT / XTT) and other mitochondrial dyes, and any combination thereof.

[0244] A10. The device, system or method of any preceding embodiment, wherein the sample comprises an agent that promotes the survival and / or division of microorganisms.

[0245] A11. The device, system, or method of any preceding embodiment, wherein one or both of the plates comprises reagents on the respective sample contact areas that promote microbial survival and / or division.

[0246] B2. The system of any preceding embodiment, further comprising:

[0247] (d) A housing configured to hold a sample and to be mounted to the mobile communication device.

[0248] B3. The system of any preceding embodiment, wherein the imager is configured to obtain a series of images of the microorganism at discrete points in time.

[0249] B4. The system of embodiment B3, wherein the processor is configured to receive and analyze the series of images, and identify and count microorganisms in a first region of each image.

[0250] B5. The system of any preceding embodiment, wherein the imager is configured to obtain a video comprising a series of images of the microorganism over a first period of time.

[0251] B6. The system of embodiment B5, wherein the processor is configured to receive and analyze the video, identify and track at least a portion of the microorganisms in the video, and evaluate changes in the tracked microorganisms over at least a portion of the first time period.

[0252] B7. The system of any preceding embodiment, wherein the housing comprises optical devices for facilitating imaging and / or signal processing of the sample by the mobile communication device, and a base configured to hold the optical devices on the mobile communication device.

[0253] B8. The system of any preceding embodiment, wherein the mobile communication device is configured to transmit the test results to medical personnel, a medical institution, or an insurance company.

[0254] B9. The system of any preceding embodiment, wherein the mobile communication device is further configured to communicate information about the subject with medical personnel, a medical institution, or an insurance company.

[0255] B10. The system of any preceding embodiment, wherein the mobile communication device is configured to receive a prescription, diagnosis, or advice from a medical professional.

[0256] B11. The system of any preceding embodiment, wherein the mobile communication device communicates with the remote location via WiFi or a cellular network.

[0257] B12. The system of any preceding embodiment, wherein the mobile communication device is a mobile phone.

[0258] B13. The system of any preceding embodiment, further comprising a temperature control element that maintains the sample deposited between the two plates at a temperature that promotes the survival and / or division of microorganisms.

[0259] C2. The method of any preceding embodiment, wherein the monitoring and de-determining steps (e) are performed by a mobile communication device comprising:

[0260] i. an imager comprising one or more cameras for imaging microorganisms in a sample; and

[0261] ii. a processor comprising electronics, a signal processor, and hardware and software for receiving and / or processing images of microorganisms and for remote communication; and

[0262] A light source from a mobile communication device or an external source, wherein the light source is configured to provide illumination to the sample for imaging with the camera.

[0263] C3. The method of any preceding embodiment, wherein the monitoring and determining step (e) comprises:

[0264] i. Obtain a series of images of platelets in a layer of uniform thickness at discrete time points;

[0265] ii. identifying and counting microorganisms in at least one region of each image; and

[0266] iii. Assessing changes in microorganisms between at least some discrete time points.

[0267] C4. The method of any preceding embodiment, wherein the monitoring and determining step (e) comprises:

[0268] i. obtaining a video comprising a series of images of the plate in a layer of uniform thickness over a first period of time;

[0269] ii. identifying and tracking at least a portion of one or more microorganisms in the video; and

[0270] iii. Assessing changes in the tracked microorganisms over at least a portion of the first time period.

[0271] C5. The method of any one of preceding embodiments C3 or C4, wherein the identifying step comprises microorganisms stained with a dye, and wherein the dye comprises a cell viability dye that indicates whether the stained cells are alive or dead.

[0272] C6. A method as described in Example C5, wherein the cell viability dye is selected from: propidium iodide, 7-AAD, trypan blue, calcein violet AM, calcein AM, fixable viability dyes, SYTO9 and other nucleic acid dyes, resazurin and colored formazan (MTT / XTT) and other mitochondrial dyes, and any combination thereof.

[0273] C7. The system of any preceding embodiment, further comprising:

[0274] When the plates are in the closed configuration, the sample is maintained at a temperature that is favorable for the survival and / or division of the microorganisms.

[0275] E1. The device, system, or method of any preceding embodiment, wherein the spacer comprises:

[0276] i. having a substantially uniform cross-section and a flat top surface of the column shape;

[0277] ii. A width-to-height ratio equal to or greater than 1;

[0278] iii. a fill factor equal to or greater than 1%; and

[0279] iv. The product of the filling factor and the Young's modulus of the spacer is 2 MPa or more.

[0280] Where fill factor is the ratio of the spacer contact area to the total board area.

[0281] E2. The apparatus, system, or method of any preceding embodiment, wherein the average value of the uniform thickness of the layer is substantially the same as the uniform height of the spacers, with a variation of less than 10%.

[0282] E3. The device, system or method of any preceding embodiment, wherein the uniform thickness layer varies by less than 30 nm.

[0283] E4. The apparatus, system, or method of any preceding embodiment, wherein the uniform thickness sample layer has a thickness uniformity of up to + / - 5%.

[0284] E5. The device, system, or method of any preceding embodiment, wherein the spacer has a column having a cross-sectional shape selected from the group consisting of: circular, polygonal, annular, square, rectangular, oval, elliptical, or any combination thereof.

[0285] E6. The device, system, or method of any preceding embodiment, wherein the spacer comprises:

[0286] i. having a substantially uniform cross-section and a flat top surface of the column shape;

[0287] ii. A width-to-height ratio equal to or greater than 1;

[0288] iii. a predetermined constant spacer spacing in the range of 10Dm to 200Dm;

[0289] iv. a fill factor equal to or greater than 1%; and

[0290] v. The product of the filling factor and the Young's modulus of the spacer is 2 MPa or more.

[0291] Where fill factor is the ratio of the spacer contact area to the total board area.

[0292] E7. An apparatus, system, or method as described in any preceding embodiment, wherein pressing the plate into a closed structure is performed in parallel or sequentially, parallel pressing applies external force on the intended area simultaneously, and sequential pressing applies external force on a portion of the intended area and gradually moves to other areas.

[0293] E8. The apparatus, system, or method of any preceding embodiment, wherein the uniform thickness sample layer has a thickness uniformity of up to + / - 5%.

[0294] Related documents

[0295] The present invention includes various embodiments that can be combined in various ways as long as the various components do not conflict with each other. The embodiments should be considered as separate invention documents: each application has the other applications as references and is also cited in its entirety for all purposes, rather than as discrete independent documents. These embodiments include not only the disclosure in the present document, but also documents cited, incorporated, or claiming priority herein.

[0296] (1) definition

[0297] The terms used to describe the devices, systems, and methods disclosed herein are defined in this application or in PCT Application Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016 and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein by reference.

[0298] (2) Spacers and uniformity

[0299] The devices, systems, and methods disclosed herein may include or use a QMAX card for sample detection, analysis, and quantification. In some embodiments, the QMAX card includes a spacer that helps to make at least a portion of the sample into a very uniform layer. The structure, materials, functions, variations, and dimensions of the spacer, as well as the uniformity of the spacer and sample layer, are disclosed herein or listed, described, and summarized in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016 and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein by reference.

[0300] (3) Hinges, notches, grooves and slides

[0301] The devices, systems, and methods disclosed herein may include or use a QMAX card for sample detection, analysis, and quantification. In some embodiments, the QMAX card includes hinges, notches, grooves, and sliders that help facilitate the operation of the QMAX card and the measurement of samples. The structures, materials, functions, variations, and dimensions of the hinges, notches, grooves, and sliders are disclosed herein or are listed, described, and summarized in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed August 10, 2016, and September 14, 2016, respectively, and in U.S. Provisional Application No. 62 / 456,065, filed February 7, 2017, all of which are incorporated herein by reference in their entirety.

[0302] (4) Card, Slider and Smartphone Inspection Systems

[0303] The devices, systems, and methods disclosed herein may include or use a QMAX card for sample detection, analysis, and quantification. In some embodiments, the QMAX card is used with a slider that allows the card to be read by a smart card detection system. The structure, materials, functions, variations, dimensions, and connections of the QMAX card, slider, and smart phone detection system are disclosed herein or listed, described, and summarized in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016, and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein by reference.

[0304] (5) Detection method

[0305] Device disclosed herein, system and method can include or be used for various types of detection methods.Detection method is disclosed herein or lists, describes and is summarized in PCT application (designated U.S.) No. PCT / US2016 / 045437 and No. PCT / US0216 / 051775, filed on August 10, 2016 and September 14, 2016, respectively, and in U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the full contents of these applications are incorporated herein by reference.

[0306] (6) Label

[0307] The devices, systems, and methods disclosed herein may use various types of tags. Tags are disclosed herein or listed, described, and summarized in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed August 10, 2016, and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed February 7, 2017, the entire contents of which are incorporated herein by reference.

[0308] (7) biomarkers

[0309] Devices, systems, and methods disclosed herein can use various types of biomarkers. Biomarkers are disclosed herein or listed, described, and summarized in PCT applications (designating the U.S.) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016 and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein by reference.

[0310] (8) Cloud

[0311] The devices, systems, and methods disclosed herein can employ cloud technologies for data transmission, storage, and / or analysis. Relevant cloud technologies are disclosed or listed, described, and summarized herein in PCT applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016, and September 14, 2016, respectively, and U.S. Provisional Application No. 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein by reference.

[0312] (9) Application (field and sample)

[0313] Device disclosed herein, system and method can be used for various applications (field and sample).Described application discloses in this article or lists, describes and is summarized in PCT application (designated U.S.) No. PCT / US2016 / 045437 and No. PCT / US0216 / 051775, submitted to respectively on August 10, 2016 and September 14, 2016, and in U.S. Provisional Application No. 62 / 456,065, submitted to on February 7, 2017, the full contents of these applications are incorporated herein by reference.

[0314] Other Notes

[0315] Other embodiments according to the inventive subject matter of the present disclosure are described in the following enumerated paragraphs.

[0316] It must be noted that, as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise, such as when the word "single" is used. For example, reference to an "analyte" includes a single analyte and multiple analytes, reference to a "capture agent" includes a single capture agent and multiple capture agents, reference to a "detection agent" includes a single detector agent and multiple detector agents, and reference to a "reagent" includes a single reagent and multiple reagents.

[0317] As used herein, the term "analyte" refers to molecules (e.g., proteins, peptides, DNA, RNA, nucleic acids or other molecules), cells, tissues, viruses and nanoparticles having different shapes. It can also be referred to as any substance suitable for testing in the present invention.

[0318] As used herein, the terms "adapted" and "configured" mean that an element, component, or other subject matter is designed and / or intended to perform a given function. Thus, the use of the terms "adapted" and "configured" should not be interpreted as meaning that a given element, component, or other subject matter is simply "capable of" performing a given function. Similarly, subject matter stated as being configured to perform a particular function may additionally or alternatively be described as being operable to perform that function.

[0319] As used herein, the phrase "for example," the phrase "as an example," and / or simply the terms "example" and "exemplary," when used with reference to one or more components, features, details, structures, embodiments, and / or methods according to the present disclosure, are intended to convey that the described components, features, details, structures, embodiments, and / or methods are illustrative, non-exclusive examples of components, features, details, structures, embodiments, and / or methods according to the present disclosure. Thus, the described components, features, details, structures, embodiments, and / or methods are not intended to be limiting, required, or exclusive / exhaustive; and other components, features, details, structures, embodiments, and / or methods, including structurally and / or functionally similar and / or equivalent components, features, details, structures, embodiments, and / or methods, are also within the scope of the present disclosure.

[0320] As used herein, the phrases "at least one" and "one or more" with respect to a list of more than one entity refer to any one or more entities in the list of entities, and are not limited to at least one of each and every entity specifically listed in the list of entities. For example, "at least one of A and B" (or equivalently, "at least one of A or B," or equivalently, "at least one of A and / or B") may refer to A alone, B alone, or A and B in combination.

[0321] As used herein, the term "and / or" placed between first and second entities means (1) the first entity, (2) the second entity, and (3) one of the first and second entities. Multiple entities listed using "and / or" should be interpreted in the same manner, i.e., "one or more" of the entities so conjoined. In addition to the entities specifically identified by the "and / or" clause, other entities may optionally be present, whether related or unrelated to those entities specifically identified.

[0322] When numerical ranges are mentioned herein, the present invention includes embodiments where the endpoints are included, embodiments where both endpoints are excluded, and embodiments where one endpoint is included but the other is excluded. Both endpoints should be assumed to be inclusive unless otherwise indicated. Furthermore, unless otherwise indicated or apparent to one of ordinary skill in the art from the context and understanding.

[0323] If any patent, patent application, or other reference is incorporated herein by reference and (1) defines a term in a manner inconsistent with the unincorporated portion of this disclosure or the other incorporated references and / or (2) is otherwise inconsistent with the unincorporated portion of this disclosure or the other incorporated references, the unincorporated portion of this disclosure shall control and the term or the disclosure incorporated therein shall control solely with respect to the reference in which the term was first defined and / or the disclosure initially appears.

[0324] 1. sample

[0325] The devices, equipment, systems and methods disclosed herein can be used for samples, such as, but not limited to, diagnostic samples, clinical samples, environmental samples and food samples. The types of samples include, but are not limited to, samples listed, described and / or summarized in PCT applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed on August 10, 2016 and September 14, 2016, respectively, the entire contents of which are incorporated herein by reference.

[0326] For example, in some embodiments, the devices, apparatus, systems, and methods disclosed herein are used with samples comprising cells, tissues, body fluids, and / or mixtures thereof. In some embodiments, the sample comprises a human body fluid. In some embodiments, the sample comprises at least one of: cells, tissues, body fluids, feces, amniotic fluid, aqueous humor, vitreous humor, blood, whole blood, fractionated blood, plasma, serum, breast milk, cerebrospinal fluid, cerumen, chyle, chyme, endolymph, perilymph, excreta, gastric acid, gastric juice, lymph, mucus, nasal drainage, sputum, pericardial fluid, peritoneal fluid, pleural fluid, pus, rheumatic fluid, saliva, sebum, semen, sputum, sweat, synovial fluid, tears, vomitus, urine, and exhaled breath condensate.

[0327] In some embodiments, the devices, apparatuses, systems and methods disclosed herein are used for environmental samples obtained from any suitable source, such as, but not limited to, liquid samples from rivers, lakes, ponds, oceans, glaciers, icebergs, rainwater, snow, sewage, reservoirs, tap water, drinking water, etc.; solid samples from soil, compost, sand, rock, concrete, wood, brick, dirt, etc.; and gaseous samples from air, underwater heat dissipation, industrial exhaust, vehicle exhaust, etc. In certain embodiments, the environmental sample is a fresh sample obtained from a source; in certain embodiments, the environmental sample is processed. For example, a sample in a non-liquid form is converted to a liquid form before applying the subject apparatus, apparatus, system and method.

[0328] In some embodiments, the devices, apparatuses, systems, and methods disclosed herein are used with food samples that are suitable or may be made suitable for animal consumption, such as human consumption. In some embodiments, food samples can include raw materials, cooked or processed foods, foods of plant and animal origin, pre-processed foods, and partially or fully processed foods. In certain embodiments, samples in non-liquid form are converted to liquid form prior to application of the subject devices, apparatuses, systems, and methods.

[0329] The devices, apparatus, systems and methods of the present invention can be used to analyze samples of any volume. Examples of volumes include, but are not limited to, about 10 mL or less, 5 mL or less, 3 mL or less, 1 microliter (μL, also herein "uL") or less, 500 μL or less, 300 μL or less, 250 μL or less, 200 μL or less, 170 μL or less, 150 μL or less, 125 μL or less, 100 μL or less, 75 μL or less, 50 μL or less, 25 μL or less, 20 μL or less, 15 μL or less, 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of these values.

[0330] In some embodiments, the volume of the sample includes but is not limited to about 100 μL or less, 75 μL or less, 50 μL or less, 25 μL or less, 20 μL or less, 15 μL or less, 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of said values. In some embodiments, the volume of the sample includes but is not limited to about 10 μL or less, 5 μL or less, 3 μL or less, 1 μL or less, 0.5 μL or less, 0.1 μL or less, 0.05 μL or less, 0.001 μL or less, 0.0005 μL or less, 0.0001 μL or less, 10 pL or less, 1 pL or less, or a range between any two of said values.

[0331] In some embodiments, the sample is of the order of a drop of liquid. In some embodiments, the sample is of the order of a drop of liquid. In some embodiments, the sample is of the order of a drop of liquid collected from a pricked finger or fingertip. In some embodiments, the sample is of the order of a drop of liquid collected from a microneedle, micropipette, or intravenous aspiration.

[0332] In certain embodiments, the sample holder is configured to hold a fluid sample. In certain embodiments, the sample holder is configured to compress at least a portion of the fluid sample into a thin layer. In certain embodiments, the sample holder includes a structure configured to heat and / or cool the sample. In certain embodiments, the heating source provides electromagnetic waves that can be absorbed by certain structures in the sample holder to change the temperature of the sample. In certain embodiments, the signal sensor is configured to detect and / or measure a signal from the sample. In certain embodiments, the signal sensor is configured to detect and / or measure an analyte in the sample. In certain embodiments, a radiator is configured to absorb heat from the sample holder and / or the heating source. In certain embodiments, the radiator includes a chamber that at least partially surrounds the sample holder.

[0333] 2 .application

[0334] The devices, apparatus, systems and methods disclosed herein can be used for various types of biological / chemical sampling, sensing, assays and applications, including PCT application (designating the United States) No. PCT / US2016 / 045437, filed on August 10, 2016, and incorporated herein by reference in its entirety.

[0335] In some embodiments, the devices, apparatuses, systems, and methods disclosed herein are used in a variety of applications across a wide range of fields where it is desirable to determine the presence or absence, quantify, and / or amplify one or more analytes in a sample. For example, in certain embodiments, the subject devices, apparatuses, systems, and methods are used to detect proteins, peptides, nucleic acids, synthetic compounds, inorganic compounds, organic compounds, bacteria, viruses, cells, tissues, nanoparticles, and other molecules, compounds, mixtures, and substances thereof. The various fields in which the subject devices, apparatuses, systems, and methods may be used include, but are not limited to, the diagnosis, management, and / or prevention of human diseases and conditions, the diagnosis, management, and / or prevention of animal diseases and conditions, the diagnosis, management, and / or prevention of plant diseases and conditions, agricultural uses, veterinary uses, food testing, environmental testing and purification, pharmaceutical testing and prevention, and the like.

[0336] Applications of the present invention include, but are not limited to: (a) detection, purification, quantification and / or amplification of compounds or biomolecules associated with certain diseases or certain stages of diseases, such as infectious and parasitic diseases, injuries, cardiovascular diseases, cancers, mental disorders, neuropsychiatric disorders and organ diseases, such as lung disease, kidney disease, (b) detection, purification, quantification and / or amplification of cells and / or microorganisms from the environment, such as viruses, fungi and bacteria, such as water, soil or biological samples, such as tissues and body fluids, (c) detection and quantification of compounds or biological samples that pose a risk to food safety, human health or national security, such as toxic waste, anthrax, (d) detection and quantification of vital parameters, such as glucose, blood oxygen level, total blood cell count, in medical or physiological monitors, (e) detection and quantification of specific DNA or RNA from biological samples (such as cells, viruses, body fluids), (f) sequencing and comparison of genetic sequences of chromosomal and mitochondrial DNA for genomic analysis, or (g) detection and quantification of reaction products, such as during the synthesis or purification of drugs.

[0337] In some embodiments, the subject devices, apparatuses, systems, and methods are used to detect nucleic acids, proteins, or other molecules or compounds in a sample. In certain embodiments, the devices, apparatuses, systems, and methods are used to rapidly, clinically detect and / or quantify one or more, two or more, or three or more disease biomarkers in a biological sample, e.g., for diagnosing, preventing, and / or managing a disease condition in a subject. In certain embodiments, the devices, apparatuses, systems, and methods are used to detect and / or quantify one or more, two or more, or three or more environmental markers in an environmental sample, e.g., a sample obtained from a river, ocean, lake, rain, snow, sewage, sewage runoff, agricultural runoff, industrial runoff, tap water, or drinking water. In certain embodiments, the devices, apparatuses, systems, and methods are used to detect and / or quantify one or more, two or more, or three or more food markers in a food sample, e.g., a sample obtained from tap water, drinking water, prepared food, processed food, or unprocessed food.

[0338] In some embodiments, the subject device is part of a microfluidic device. In some embodiments, the subject devices, apparatuses, systems, and methods are used to detect fluorescent or luminescent signals. In some embodiments, the subject devices, apparatuses, systems, and methods include or are used with a communication device, such as, but not limited to, a mobile phone, a tablet computer, and a portable computer. In some embodiments, the subject devices, apparatuses, systems, and methods include or are used with an identifier, such as, but not limited to, an optical barcode, a radio frequency ID tag, or a combination thereof.

[0339] In some embodiments, the sample is a diagnostic sample obtained from a subject, the analyte is a biomarker, and the amount of the analyte measured in the sample can be diagnostic of a disease or condition. In some embodiments, the subject devices, systems, and methods further comprise receiving or providing a report to the subject indicating the measured amount of the biomarker and a range of measured values ​​for the biomarker in an individual who does not have or is at low risk of having the disease or condition, wherein the measured amount of the biomarker relative to the range of measured values ​​is used to diagnose the disease or condition.

[0340] In some embodiments, the sample is an environmental sample, and wherein the analyte is an environmental marker. In some embodiments, the subject device, system and method include receiving or providing a report indicating the safety or harmfulness of the subject exposed to the environment from which the sample was obtained. In some embodiments, the subject device, system and method include sending data comprising the amount of the measured environmental marker to a remote location and receiving a report indicating the safety or harmfulness of the subject exposed to the environment from which the sample was obtained.

[0341] In some embodiments, the sample is a food sample, wherein the analyte is a food marker, and wherein the amount of the food marker in the sample is correlated with the safety of consuming the food. In some embodiments, the subject devices, systems, and methods include receiving or providing a report indicating the safety or hazardousness of consuming the food from which the sample was obtained by the subject. In some embodiments, the subject devices, systems, and methods include transmitting data containing a measured amount of the food marker to a remote location and receiving a report indicating the safety or hazardousness of consuming the food from which the sample was obtained by the subject.

[0342] 3. Analytes, Biomarkers, and Diseases

[0343] The devices, apparatus, systems, and methods disclosed herein can be used for the detection, purification, and / or quantification of various analytes. In some embodiments, the analytes are biomarkers associated with various diseases. In some embodiments, the analytes and / or biomarkers indicate the presence, severity, and / or stage of a disease. Analytes, biomarkers and / or diseases that can be detected and / or measured using the devices, apparatus, systems and / or methods of the present invention include those listed, described and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 045437, filed on August 10, 2016, PCT / US2016 / 054025, filed on September 27, 2016, and U.S. Provisional Application Nos. 62 / 234,538, filed on September 29, 2015, 62 / 233,885, filed on September 28, 2015, 62 / 293,188, filed on February 9, 2016, and 62 / 305,123, filed on March 8, 2016, which are hereby incorporated by reference in their entireties. For example, the devices, apparatus, systems and methods disclosed herein can be used for (a) detection, purification and quantification of compounds or biomolecules associated with certain disease stages, such as infectious and parasitic diseases, injuries, cardiovascular diseases, cancers, mental disorders, neuropsychiatric disorders and organic diseases, such as lung disease and kidney disease, (b) detection, purification and quantification of microorganisms, such as viruses, fungi and bacteria, from the environment, such as water, soil or biological samples, such as tissues and body fluids, (c) detection and quantification of chemical compounds or biological samples that pose a threat to food safety or national security, such as toxic waste, anthrax, (d) quantification of vital parameters, such as glucose, blood oxygen levels, total blood cell counts, in medical or physiological monitors, (e) detection and quantification of specific DNA or RNA from biological samples, such as cells, viruses, body fluids, (f) sequencing and comparison of genetic sequences of chromosomal and mitochondrial DNA for genomic analysis, or (g) detection of reaction products, such as during drug synthesis or purification.

[0344] In some embodiments, the analyte can be a biomarker, an environmental marker, or a food marker. In some cases, the sample is a liquid sample and can be a diagnostic sample (e.g., saliva, serum, blood, sputum, urine, sweat, tears, semen, or mucus); an environmental sample obtained from a river, ocean, lake, rainwater, snow, sewage, sewage runoff, agricultural runoff, industrial runoff, tap water, or drinking water; or a food sample obtained from tap water, drinking water, prepared food, processed food, or unprocessed food.

[0345] In any embodiment, the sample can be a diagnostic sample obtained from a subject, the analyte can be a biomarker, and the amount of the analyte in the sample measured can be diagnostic of a disease or condition.

[0346] In any embodiment, the devices, apparatus, systems and methods of the present invention may further include diagnosing the subject based on information including the measured amount of the biomarker in the sample. In some cases, the diagnosing step includes sending data including the measured amount of the biomarker to a remote location and receiving a diagnosis based on the information including the measurement from the remote location.

[0347] In any embodiment, the biomarker can be selected from Table B1, 2, 3 or 7 disclosed in U.S. Provisional Application Nos. 62 / 234,538, 62 / 293,188 and / or 62 / 305,123 and / or PCT Application No. PCT / US2016 / 054,025, the entire contents of which are incorporated herein for all purposes. In some cases, the biomarker is a protein selected from Table B1, 2 or 3. In some cases, the biomarker is a nucleic acid selected from Table B2, 3 or 7. In some cases, the biomarker is an infectious agent-derived biomarker selected from Table B2. In some cases, the biomarker is a microRNA (miRNA) selected from Table B7.

[0348] In any embodiment, applying step b) can include isolating miRNA from the sample to produce an isolated miRNA sample, and applying the isolated miRNA sample to a disk-coupled dot-rod antenna (QMAX device) array.

[0349] In any embodiment, the QMAX device can contain a plurality of capture agents that each bind a biomarker selected from Tables B1, B2, B3, and / or B7, wherein the reading step d) comprises obtaining a measurement of the amount of the plurality of biomarkers in the sample, and wherein the amount of the plurality of biomarkers in the sample is diagnostic for a disease or condition.

[0350] In any embodiment, the capture agent can be an antibody epitope, and the biomarker can be an antibody that binds to the antibody epitope. In some embodiments, the antibody epitope comprises a biomolecule or fragment thereof selected from Table B4, B5, or B6. In some embodiments, the antibody epitope comprises an allergen or fragment thereof selected from Table B5. In some embodiments, the antibody epitope comprises a biomolecule or fragment thereof derived from an infectious agent selected from Table B6.

[0351] In any embodiment, the QMAX device can contain a plurality of antibody epitopes selected from Tables B4, B5, and / or B6, wherein reading step d) comprises obtaining a measurement of the amount of the plurality of epitope-binding antibodies in the sample, and wherein the amount of the plurality of epitope-binding antibodies in the sample is diagnostic for the disease or condition.

[0352] In any embodiment, the sample can be an environmental sample, and wherein the analyte can be an environmental marker. In some embodiments, the environmental marker is selected from Table B8 in U.S. Provisional Application No. 62 / 234,538 and / or PCT Application No. PCT / US2016 / 054025.

[0353] In any embodiment, the method can include receiving or providing a report indicating the safety or harmfulness to the subject of exposure to the environment from which the sample was obtained.

[0354] In any embodiment, the method can include transmitting data comprising the measured amount of the environmental marker to a remote location and receiving a report indicating the safety or hazardousness to the subject of exposure to the environment from which the sample was obtained.

[0355] In any embodiment, the QMAX device array can include a plurality of capture agents, each capture agent binding an environmental marker selected from Table B8, and wherein the reading step d) can include obtaining a measurement of the amount of the plurality of environmental markers in the sample.

[0356] In any embodiment, the sample can be a food sample, wherein the analyte can be a food marker, and wherein the amount of the food marker in the sample can be correlated with the safety of the consumed food. In some embodiments, the food marker is selected from Table B9.

[0357] In any embodiment, the method can include receiving or providing a report indicating the safety or hazards of consuming by the subject a food from which the sample was obtained.

[0358] In any embodiment, the method can include transmitting data containing the measured amounts of the food markers to a remote location and receiving a report indicating the safety or harmfulness of the subject consuming the food from which the sample was obtained.

[0359] In any embodiments, the devices, apparatus, systems, and methods disclosed herein may comprise a plurality of capture agents that each bind a food marker selected from Table B9, Table B9 from U.S. Provisional Application No. 62 / 234,538 and PCT Application No. PCT / US2016 / 054025, wherein the obtaining may comprise obtaining a measurement of the amount of the plurality of food markers in the sample, and wherein the amount of the plurality of food markers in the sample may be correlated to the safety of the food to be consumed.

[0360] Also provided herein are kits for practicing the devices, systems, and methods of the present invention.

[0361] The sample size can be about one drop of sample. The sample size can be the amount collected from a finger prick or puncture. The sample size can be the amount collected from a microneedle or intravenous aspiration.

[0362] The sample can be used without further processing after being obtained from the source, or can be processed, for example, to enrich for an analyte of interest, remove large particulate matter, dissolve or resuspend a solid sample, etc.

[0363] Any suitable method for applying the sample to the QMAX device can be used. Suitable methods may include using a pipette, dropper, syringe, etc. In certain embodiments, when the QMAX device is on a holder in the form of a dipstick, as described below, the sample can be applied to the QMAX device by dipping the sample receiving area of ​​the dipstick into the sample.

[0364] The sample can be collected one or more times. Samples collected over time can be aggregated and / or processed individually (as described herein, by applying to a QMAX device and obtaining a measurement of the amount of analyte in the sample). In some cases, the measurements obtained over time can be aggregated and used for longitudinal analysis over time to facilitate screening, diagnosis, treatment, and / or disease prevention.

[0365] Washing the QMAX device to remove unbound sample components can be performed in any convenient manner, as described above. In certain embodiments, the surface of the QMAX device is washed with a binding buffer to remove unbound sample components.

[0366] Detectable labeling of the analyte can be performed by any convenient method. The analyte can be labeled directly or indirectly. In direct labeling, the analyte in the sample is labeled before the sample is applied to the QMAX device. In indirect labeling, unlabeled analytes in the sample are labeled after the sample is applied to the QMAX device to capture the unlabeled analyte, as described below.

[0367] 4. Label

[0368] The devices, apparatus, systems, and methods disclosed herein can be used with various types of tags, including tags disclosed, described, and / or summarized in PCT Application No. (designating the United States) PCT / US2016 / 045437, filed on August 10, 2016, and incorporated herein by reference in its entirety.

[0369] In some embodiments, the label is optically detectable, such as, but not limited to, a fluorescent label. In some embodiments, the label includes, but is not limited to, IRDye 800CW, Alexa 790, Dylight 800, fluorescein, fluorescein isothiocyanate, succinimidyl ester of carboxyfluorescein, succinimidyl ester of fluorescein, 5-isomer of fluorescein dichlorotriazine, caged carboxyfluorescein-alanine-carboxamide, Oregon Green 488, Oregon Green 514; fluorescein yellow, acridine orange, rhodamine, tetramethylrhodamine, Texas Red, propidium iodide, JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide), tetrabromorhodamine 123, rhodamine 6G, TMRM (tetramethylrhodamine methyl ester), TMRE (tetramethylrhodamine ethyl ester), tetramethylrhodamine, rhodamine B, and 4-dimethylaminotetramethylrhodamine. Ming, green fluorescent protein, blue-shifted green fluorescent protein, blue-green-shifted green fluorescent protein, red-shifted green fluorescent protein, yellow-shifted green fluorescent protein, 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid; acridine and derivatives, for example, acridine, acridine isothiocyanate; 5-(2'-aminoethyl)aminonaphthalene-1-sulfonic acid (EDANS); 4-amino-N-[3-vinylsulfonyl)phenyl]naphthamide-3,5-disulfonate; N-(4-anilino-1-naphthyl)maleimide; anthranilamide; 4,4-difluoro-5-(2-thienyl)-4-boron-3a,4a-diaza-5-indacene-3-propionic acid BODIPY; grade Blue; brilliant yellow; coumarin and derivatives: coumarins, 7-amino-4-methylcoumarin (AMC, coumarin 120), 7-amino-4-methylcoumarin (AMC, coumarin 151); cyanine dyes; cyanosine; 4',6-diamino-2-phenylindole (DAPI); 5',5"-dibromopyrogallolsulfonaphthalene (bromopyrogallol red); 7-diethylamino-3-(4'-isothiocyanatophenyl)-4-methylcoumarin; diethylenetriamine pentaacetate; 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid; 4,4'-diisothiocyanato-2,2'-yldisulfonic acid disodium salt; 5-(dimethylamino)naphthalene-1-sulfonyl Chloride (DNS, dansyl chloride); 4-dimethylaminophenylazophenyl-4'-isothiocyanate (DABITC); Eosin and derivatives: Eosin, Eosin isothiocyanate, Erythrosine and derivatives: Erythrosine B, Erythrosine, isothiocyanate; Ethidium; Fluorescein and its derivatives: 5-carboxyfluorescein (FAM), 5-(4,6-dichlorotriazin-2-yl)amino-fluorescein (DTAF), 2',7'-dimethoxy-4',5'-dichloro-6-carboxyfluorescein (JOE), Fluorescein, Fluorescein isothiocyanate, QFITC, (XRITC); Fluorescein amine; IR144; IR1446; Malachite green isothiocyanate; 4-Methylumbelliferyl o-cresolphthalein;Nitrotyrosine; para-carmine; phenol red; B-phycoerythrin; o-phthalaldehyde; pyrene and its derivatives: pyrene, pyrenebutyric acid, succinimidyl 1-pyrene; butyrate quantum dots; Reactive Red 4 (Cibacron; TM Brilliant Red 3B-A) Rhodamine and derivatives: 6-carboxy-X-rhodamine (ROX), 6-carboxyrhodamine (R6G), lissamine rhodamine B sulfonyl chloride rhodamine (Rhod), Rhodamine B, Rhodamine 123, Rhodamine X isothiocyanate, sulforhodamine B, sulforhodamine 101, sulfonyl chloride derivatives of 5-sulforhodamine (Texas Red); N,N,N',N'-tetramethyl-6-carboxyrhodamine TAMRA; tetramethylrhodamine; tetramethylrhodamine isothiocyanate (TRITC); riboflavin; 5-(2'-aminoethyl)aminonaphthalene-1-sulfonic acid (EDANS), 4-(4'-dimethylaminophenylazo)benzoic acid (DABCYL), rosocyanic acid; CAL Fluorescent Orange 560; terbium chelate derivatives; Cy3; Cy5; Cy5.5; Cy7; IRD700; IRD 800; La Jolla Blue; phthalocyanines; and naphthalocyanines, coumarins and related dyes, xanthene dyes, such as rhodols, resorufins, bimanes, acridines, isoindoles, dansyl dyes, aminophthalhydrazides (e.g., luminol), and isoluminol derivatives, aminophthalimides, aminonaphthalimides, aminobenzofurans, aminoquinolines, dicyanohydroquinones, fluorescent europium and terbium complexes; combinations thereof, and the like. Suitable fluorescent and chromogenic proteins include, but are not limited to, green fluorescent protein (GFP), including, but not limited to, GFP from Aequoria victoria or a derivative thereof, e.g., a "humanized" derivative, such as enhanced GFP; GFP from another species, e.g., Renilla reniformis, Renilla mulleri, or Ptilosarcus guernyi; "humanized" recombinant GFP (hrGFP); any of a variety of fluorescent and colored proteins from coral species; any combination thereof; and the like.

[0370] 5. QMAX device

[0371] The devices, apparatus, systems and methods disclosed herein may include or use a QMAX device ((Q: quantification; M: amplification; A: addition of reagents; X: acceleration; also referred to in some embodiments as a Q card or compression regulated open flow (CROF) device), including the QMAX devices listed, described and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 045437 filed on August 10, 2016 and U.S. Provisional Application No. 62,431,639 filed on December 9, 2016 and No. 62 / 456,287 filed on February 8, 2017, the entire contents of which are incorporated herein by reference.

[0372] As used herein, the terms "CROF card (or card)", "COF card", "QMAX card", "Q card", "CROF device", "COF device", "QMAX device", "CROF board", "COF board", and "QMAX board" are interchangeable, except that in some embodiments, the COF card does not include a spacer; and these terms refer to a device that includes a first board and a second board that are movable relative to each other into different configurations (including an open configuration and a closed configuration) and that includes a spacer that adjusts the spacing between the boards (except in some embodiments of the COF). The term "X-board" refers to one of the two boards in a CROF card, with a spacer fixed to the board. Further description of COF cards, CROF cards, and X-boards is described in provisional application serial number 62 / 456,065, filed on February 7, 2017, the entire contents of which are incorporated herein for all purposes.

[0373] The term "Compression Open Flow (COF)" refers to a method of changing the shape of a flowable sample deposited on plates by (i) placing another plate on top of at least a portion of the sample, and (ii) then compressing the sample between two plates by pushing the two plates toward each other; wherein the compression reduces the thickness of at least a portion of the sample and causes the sample to flow into the open space between the plates. The term "Compression Regulated Open Flow" or "CROF" (or "Self-Calibrated Compression Open Flow" or "SCOF" or "SCCOF") (also known as QMAX) refers to a specific type of COF in which the final thickness of part or all of the sample after compression is "regulated" by a spacer, which is placed between the two plates. Herein, CROF device is used interchangeably with QMAX card.

[0374] In the QMAX process, the term "open configuration" of two plates refers to a structure in which the two plates are either partially or completely separated and the spacing between the plates is not adjusted by spacers.

[0375] The term "closed configuration" of two plates in QMAX processing refers to a configuration in which the plates face each other, the spacer and the associated volume of the sample are between the plates, the associated spacing between the plates and therefore the thickness of the associated volume of the sample are adjusted by the plates and the spacer, wherein the associated volume is at least a portion of the entire volume of the sample.

[0376] In the QMAX process, the term "sample thickness is adjusted by the plate and spacer" means that for given conditions of the plate, sample, spacer and plate compression method, the thickness of at least one port of the sample in the closed configuration of the plate can be predetermined based on the properties of the spacer and the plate.

[0377] The term "inner surface" or "sample surface" of the plate in the QMAX card refers to the surface of the plate that contacts the sample, while the other surface of the plate (not contacting the sample) is called the "outer surface."

[0378] Unless otherwise specified, the terms "height" or "thickness" of an object in the QMAX process refer to the dimension of the object in the direction perpendicular to the plate surface. For example, the spacer height is the dimension of the spacer in the direction perpendicular to the plate surface, and the spacer height and spacer thickness mean the same thing.

[0379] Unless otherwise specified, the term "area" of an object in the QMAX process refers to the area of ​​the object parallel to the plate surface. For example, a spacer area is the area of ​​the spacer parallel to the plate surface.

[0380] The term QMAX card refers to a device that performs a QMAX (eg, CROF) process on a sample, and may or may not have a hinge connecting the two plates.

[0381] The terms "QMAX card with hinge" and "QMAX card" are interchangeable.

[0382] The terms "angle self-maintaining," "angle self-maintaining," or "rotation angle self-maintaining" refer to a property of a hinge that substantially maintains the angle between two panels after the external force moving the panels from the initial angle to the angle is removed from the panels.

[0383] When using a QMAX card, both plates need to be opened first for sample deposition. However, in some embodiments, the QMAX card from the package has both plates in contact with each other (e.g., closed position), and because one or both plates are very thin, separating them can be challenging. To facilitate opening of the QMAX card, one or more opening notches are created at or near an edge or corner of the first plate, and in the closed position of the plates, a portion of the second plate is placed over the opening notches, so that the second plate can be lifted open in the notch of the first plate without blocking the first plate.

[0384] In the QMAX assay platform, the QMAX card uses two plates to manipulate the sample into a thin layer (e.g., by compressing it). In some embodiments, plate manipulation requires repeated changes in the relative position of the two plates (referred to as plate configuration) by manual or other external forces. The QMAX card is designed to facilitate easy and rapid manual manipulation.

[0385] In the QMAX assay, one of the plate configurations is an open configuration, in which the two plates are fully or partially separated (the spacing between the plates is not controlled by a spacer) and the sample can be deposited. The other configuration is a closed configuration, in which at least a portion of the sample deposited in the open configuration is compressed by the two plates into a layer of very uniform thickness, the uniform thickness of the layer being defined by the inner surfaces of the plates and adjusted by the plates and spacers. In some embodiments, the average spacing between the two plates is greater than 300 μm.

[0386] In the QMAX assay operation, the operator needs to first place both plates in an open configuration ready for sample deposition, then deposit the sample on one or both plates, and finally close the plates to a closed position. In certain embodiments, the two plates of the QMAX card are initially on top of each other and need to be separated to enter the open configuration for sample deposition. When one of the plates is a thin plastic film (175 μm thick PMA), this separation is difficult to perform manually. The present invention is intended to provide devices and methods that make the operation of certain assays (such as the QMAX card assay) easy and rapid.

[0387] In some embodiments, a QMAX device includes a hinge that connects two or more panels together, allowing the panels to open and close in a manner similar to a book. In some embodiments, the hinge's material allows the hinge to self-maintain the angle between the panels after adjustment. In some embodiments, the hinge is configured to hold the QMAX card in a closed configuration, allowing the entire QMAX card to slide in and out of the card slot without accidentally separating the two panels. In some embodiments, the QMAX device includes one or more hinges capable of controlling the rotation of more than two panels.

[0388] In some embodiments, the hinge is made of a metal material selected from gold, silver, copper, aluminum, iron, tin, platinum, nickel, cobalt, an alloy, or any combination thereof. In some embodiments, the hinge comprises a single layer made of a polymer material, such as, but not limited to, plastic. The polymer material is selected from the group consisting of: acrylate polymers, vinyl polymers, olefin polymers, cellulosic polymers, non-cellulosic polymers, polyester polymers, nylon, cyclic olefin copolymer (COC), poly(methyl methacrylate) (PMMB), polycarbonate (PC), cyclic olefin polymer (COP), liquid crystal polymer (LCP), polyamide (PB), polyethylene (PE), polyimide (PI), polypropylene (PP), polyphenylene ether (PPE), polystyrene (PS), polyoxymethylene (POM), polyetheretherketone (PEEK), polyethersulfone (PES), polyethylene phthalate (PET), polytetrafluoroethylene (PTFE), polyvinyl chloride (PVC), polyvinylidene fluoride (PVDF), polybutylene terephthalate (PBT), fluorinated ethylene propylene (FEP), perfluoroalkoxyalkane (PFB), polydimethylsiloxane (PDMS), rubber, or any combination thereof. In some embodiments, the polymer material is selected from polystyrene, PMMB, PC, COC, COP, other plastics, or any combination thereof.

[0389] In some embodiments, the QMAX device includes an opening mechanism, such as but not limited to notches on the edge of the panel or strips attached to the panel, making it easier for the user to manipulate the positioning of the panel, such as but not limited to separating the panels by hand.

[0390] In some embodiments, the QMAX device comprises grooves on one or both plates. In certain embodiments, the grooves restrict the flow of sample on the plates.

[0391] 6. spacers

[0392] The devices, apparatus, systems, and methods disclosed herein may include or use a device (e.g., a QMAX device) that includes spacers listed, described, and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 045437, filed August 10, 2016, and U.S. Provisional Application Nos. 62,431,639, filed December 9, 2016, and 62 / 456,287, filed February 8, 2017, the entire contents of which are incorporated herein by reference.

[0393] Essentially, unless otherwise specified, the term "spacer" or "stopper" refers to a mechanical object that, when placed between two plates, sets a limit on the minimum spacing between the two plates that can be reached when the two plates are compressed together. That is, during compression, the spacer will stop the relative motion of the two plates to prevent the plate spacing from becoming less than a preset (i.e., predetermined) value.

[0394] The terms "spacers having a predetermined height" and "spacers having a predetermined spacer spacing" respectively mean that the values ​​of the spacer height and the spacer spacing are known prior to the QMAX process. If the values ​​of the spacer height and the spacer spacing are not known prior to the QMAX process, then the values ​​of the spacer height and the spacer spacing are not predetermined. For example, in the case of beads being sprayed onto a plate as spacers, where the beads land at random locations on the plate, the distance between the spacers is not predetermined. Another example of an unpredictable inter-spacer distance is when the spacers move during the QMAX process.

[0395] In the context of the QMAX process, the term "a spacer is fixed to its corresponding plate" means that the spacer is attached to a position on the plate and remains attached to that position during the QMAX process (i.e., the position of the spacer on the corresponding plate does not change). An example of a "spacer being fixed to its corresponding plate" is when the spacer is integrally made from one piece of material from the plate and the position of the spacer relative to the plate surface does not change during the QMAX process. An example of a "spacer being not fixed to its corresponding plate" is when the spacer is bonded to the plate by an adhesive, but during use of the plate, during the QMAX process, the adhesive fails to hold the spacer in its original position on the plate surface and the spacer moves away from its original position on the plate surface.

[0396] 7. adapter

[0397] The devices, apparatus, systems, and methods disclosed herein can be used with an adapter configured to accommodate the device and connect the device to a reader, such as, but not limited to, a smartphone. In some embodiments, a Q card is used with a slider that allows the card to be inserted into the adapter so that the card can be read by a smartphone detection system. The structure, materials, functions, variations, dimensions, and connections of the Q card, slider, and adapter are disclosed, listed, described, and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 045437, filed August 10, 2016, and No. PCT / US0216 / 051775, filed September 14, 2016, U.S. Provisional Application No. 62 / 456,590, filed February 8, 2017, No. 62 / 459,554, filed February 15, 2017, and No. 62 / 460,075, filed February 8, 2017, all of which are incorporated herein in their entirety for all purposes.

[0398] In some embodiments, the adapter includes a receptacle slot configured to receive the QMAX device when the device is in a closed configuration. In some embodiments, the QMAX device has a sample stored therein, and the adapter can be connected to a mobile device (e.g., a smartphone) so that the sample can be read by the mobile device. In some embodiments, the mobile device can detect and / or analyze a signal from the sample. In some embodiments, the mobile device can capture an image of the sample while the sample is in the QMAX device and within the field of view (FOV) of a camera, which in some embodiments is part of the mobile device.

[0399] In some embodiments, the adapter includes a plurality of optical components configured to enhance, amplify, and / or optimize the generation of a signal from the sample. In some embodiments, the optical components include a plurality of sections configured to enhance, amplify, and / or optimize the illumination provided to the sample. In certain embodiments, the illumination is provided by a light source that is part of the mobile device. In some embodiments, the optical components include a plurality of sections configured to enhance, amplify, and / or optimize the signal from the sample. In some embodiments, the structure, function, and construction of the optical components can be found in PCT Application (designating the United States) No. PCT / US2016 / 045437, filed on August 10, 2016, and No. PCT / US0216 / 051775, filed on September 14, 2016, U.S. Provisional Application No. 62 / 456,590, filed on February 8, 2017, No. 62 / 459,554, filed on February 15, 2017, and No. 62 / 460,075, filed on February 8, 2017, all of which are incorporated herein in their entirety for all purposes.

[0400] 8. size

[0401] The devices, apparatuses, systems, and methods disclosed herein may include or utilize a QMAX device, which may include a plate and a spacer. In some embodiments, the dimensions of the various components of the QMAX device and its adapters are listed, described, and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 045437, filed August 10, 2016, and U.S. Provisional Application Nos. 62,431,639, filed December 9, 2016, and 62 / 456,287, filed February 8, 2017, which are incorporated herein by reference in their entirety.

[0402] In some embodiments, the dimensions are listed in the following table:

[0403] plate:

[0404]

[0405]

[0406] Hinge:

[0407]

[0408]

[0409] Notch:

[0410]

[0411] Grooves:

[0412]

[0413]

[0414] socket slot

[0415]

[0416] 9. Manual pressing

[0417] For the devices, apparatus, systems and methods disclosed herein, human hands can be used to manipulate or operate the plates and / or samples. In some embodiments, human hands can be used to press the plates into a closed configuration; in some embodiments, human hands can be used to press the samples into a thin layer. Methods of using hand pressure are described and / or summarized in PCT Application (designating the United States) No. PCT / US2016 / 046437 filed on August 10, 2016 and PCT / US2016 / 051775 filed on September 14, 2016, and U.S. Provisional Application No. 62 / 431,639 filed on December 9, 2016, No. 62 / 456,287 filed on February 8, 2017, No. 62 / 456,065 filed on February 7, 2017, No. 62 / 456,504 filed on February 8, 2017, and No. 62 / 460,062 filed on February 16, 2017, all of which are incorporated herein by reference.

[0418] In some embodiments, the plates of the QMAX device can be manipulated or operated by a human hand. In certain embodiments, a human hand can be used to apply an imprecise force to compress the plates from an open configuration to a closed configuration. In certain embodiments, a human hand can be used to apply an imprecise force to achieve a high level of uniformity in sample thickness (e.g., less than 5%, 10%, 15%, or 20% variability).

[0419] In certain embodiments, the plates are movable relative to each other into different configurations, including an open configuration and a closed configuration; in the open configuration, the two plates are partially or completely separated, the spacing between the plates is not adjusted by spacers, and the sample is deposited on one or both of the plates; in the closed configuration: at least a portion of the sample is compressed by the two plates into a layer of very uniform thickness and is essentially stationary relative to the plates, wherein the uniform thickness of the layer is defined by the sample contact area of ​​the two plates and adjusted by the plates and spacers; in some embodiments, the force pressing the two plates into the closed configuration is an imprecise pressing force provided by a human hand.

[0420] In some embodiments, the panels are conformally pressed. In some embodiments, at least one region of the panels is conformally pressed in parallel or sequentially to press the panels together into a closed configuration, wherein conformal pressing produces substantially uniform pressure on at least a portion of the sample of the panels, and the pressing causes at least a portion of the sample to expand laterally between the sample-contacting surfaces of the panels, and wherein the closed configuration is a configuration in which the spacing between panels in a layer of uniform thickness regions is adjusted by spacers; in some embodiments, conformal pressing is a method of making the pressure applied to an area substantially constant regardless of changes in the shape of the outer surface of the panels. In some embodiments, parallel pressing applies pressure to the intended regions simultaneously, and sequential pressing applies pressure to a portion of the intended region and gradually moves to other regions.

[0421] In some embodiments, the panels are pressed into the closed configuration by an imprecise force. In certain embodiments, the imprecise force is applied by a human hand. In some embodiments, the pressure is an imprecise force having a magnitude that is (a) unknown and unpredictable at the time the force is applied, or (b) unknown and cannot be predicted within an accuracy of 30% or better than the applied force. In some embodiments, the pressure is an imprecise force having a magnitude that cannot be determined at the time the force is applied to within an accuracy of 30%, 40%, 50%, 70%, 100%, 200%, 300%, 500%, 1000%, 2000%, or any range therebetween.

[0422] 10. smartphone

[0423] The devices, apparatus, systems, and methods disclosed herein can be used with mobile devices such as, but not limited to, smartphones. Smartphone detection techniques are disclosed herein or listed, described, and summarized in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed August 10, 2016, and September 14, 2016, respectively; U.S. Provisional Application No. 62 / 456,065, filed February 7, 2017; U.S. Provisional Application No. 62 / 456,287, filed February 8, 2017; and U.S. Provisional Application No. 62 / 456,504, filed February 8, 2017, all of which are incorporated herein in their entirety for all purposes.

[0424] In some embodiments, the smartphone includes a camera that can be used to capture an image or sample when the sample is in the camera's field of view (e.g., via an adapter). In some embodiments, the camera includes a single set of lenses (e.g., an iPhone™ 6). In some embodiments, the camera includes at least two sets of lenses (e.g., an iPhone™ 7). In some embodiments, the smartphone includes a camera, but the camera is not used for image capture.

[0425] In some embodiments, the smartphone includes a light source, such as, but not limited to, an LED (light emitting diode). In certain embodiments, the light source is used to illuminate the sample when the sample is in the field of view of the camera (e.g., via an adapter). In some embodiments, the light from the light source is enhanced, amplified, modified, and / or optimized by the optical components of the adapter.

[0426] In some embodiments, the smartphone includes a processor configured to process information from the sample. The smartphone includes software instructions that, when executed by the processor, can enhance, amplify, and / or optimize the signal (e.g., image) from the sample. The processor may include one or more hardware components, such as a central processing unit (CPU), an application specific integrated circuit (ASIC), an application specific instruction set processor (ASIP), a graphics processing unit (GPU), a physical processing unit (PPU), a digital signal processor (DSP), a field programmable gate array (FPGA), a programmable logic device (PLD), a controller, a microcontroller unit, a reduced instruction set computer (RISC), a microprocessor, etc., or any combination thereof.

[0427] In some embodiments, the smartphone includes a communication unit configured and / or used to transmit data and / or images related to the sample to another device. By way of example only, the communication unit can use a cable network, a wired network, an optical fiber network, a telecommunications network, an intranet, the Internet, a local area network (LAN), a wide area network (WAN), a wireless local area network (WLAN), a metropolitan area network (MAN), a wide area network (WAN), a public switched telephone network (PSTN), a Bluetooth network, a ZigBee network, a near field communication (NFC) network, etc., or any combination thereof.

[0428] In some embodiments, the smartphone is an iPhone™, an Android™ phone, or a Windows™ phone.

[0429] 11. Cloud

[0430] The devices, apparatus, systems, and methods disclosed herein can be used with cloud storage and computing technologies. Related cloud technologies are disclosed or listed, described, and summarized herein in PCT Applications (designating the United States) Nos. PCT / US2016 / 045437 and PCT / US0216 / 051775, filed August 10, 2016, and September 14, 2016, respectively; U.S. Provisional Application No. 62 / 456,065, filed February 7, 2017; U.S. Provisional Application No. 62 / 456,287, filed February 8, 2017; and U.S. Provisional Application No. 62 / 456,504, filed February 8, 2017, all of which are incorporated herein in their entirety for all purposes.

[0431] In some embodiments, cloud storage and computing technology may involve a cloud database. By way of example only, a cloud platform may include a private cloud, a public cloud, a hybrid cloud, a community cloud, a distributed cloud, an inter-cloud, a multi-cloud, or any combination thereof. In some embodiments, a mobile device (e.g., a smartphone) may connect to the cloud via any type of network, including a local area network (LAN) or a wide area network (WAN).

[0432] In some embodiments, data associated with the sample (e.g., an image of the sample) is sent to the cloud without processing by the mobile device and can be further analyzed remotely. In some embodiments, data associated with the sample is processed by the mobile device and the results are sent to the cloud. In some embodiments, both the raw data and the results are sent to the cloud.

Claims

1. A device for examining the effect of a test agent on one or more individual microorganisms in a sample, comprising: a first plate, a second plate, the test reagent, the sample, and the detection reagent being placed between the two plates, wherein the test reagent is a fluorescent dye for labeling individual microorganisms, and one or both plates have a spacer; is squeezed between the plates to form a uniform test layer; an imager for acquiring images of signals emanating from said individual microorganisms in said uniform test layer at at least two discrete points in time; a processor for analyzing the images at the two discrete time points to determine changes in the individual microorganisms between the two discrete time points; The test reagent and the sample containing one or more microorganisms are placed between two plates to form a uniform test layer; wherein the two plates are movable relative to each other into different configurations, including an open configuration and a closed configuration; wherein in the open configuration, the two plates are partially or completely separated, the spacing between the plates is not adjusted by the spacer, and the sample is deposited on one or both of the plates; as well as wherein in the closed configuration, which is configured after the sample is deposited in the open configuration: at least a portion of the sample is compressed by the two plates into the uniform test layer having a substantially uniform thickness, and the uniform thickness of the layer is defined by the sample contacting surfaces of the plates and adjusted by the plates and the spacers; wherein said imaging is performed in a closed configuration, and wherein in said closed configuration said test layer thickness is configured such that there is substantially no overlap between any two individual microorganisms; and Wherein in the closed configuration, the thickness of the uniform test layer is adjusted by the two plates and the spacers in the sample area, and the height of the spacers is 250 microns or less.

2. A device for examining the effect of a test agent on one or more individual microorganisms in a sample, comprising: a first plate, a second plate, wherein the test reagent and the sample containing one or more microorganisms are placed between the two plates to form a uniform test layer; an imager for acquiring at least two images of the uniform test layer at discrete points in time; a processor for analyzing the images to determine changes in the individual microorganisms between at least two of the discrete time points; wherein the thickness of the uniform test layer is adjusted by two plates and spacers between the two plates and within the sample; wherein the thickness of the test layer is configured such that there is substantially no overlap between two microorganism individuals; wherein the two plates are movable relative to each other into different configurations, including an open configuration and a closed configuration; wherein in the developed configuration, the two plates are partially or fully separated, the spacing between the two plates is not regulated by the spacer, and the sample is deposited on one or both plates; as well as wherein in the closed configuration, the closed configuration is configured after the sample is deposited in the open configuration; at least a portion of the sample is compressed by the two plates into the uniform test layer having a very uniform thickness, and the uniform thickness of the layer is defined by the sample contact surface of the plates and adjusted by the plates and the spacers.

3. The apparatus according to claim 1 or 2, wherein the imager comprises a camera or a video camera configured to obtain a series of images of the microorganism at discrete time points.

4. The apparatus of claim 1 or 2, wherein the imager is configured to obtain a video comprising a series of images of the microorganism over a first time period.

5. The device according to claim 1 or 2, wherein the uniform height of the spacers is equal to or less than the average size of the one or more microorganisms; or The uniform height of the spacers is in the range of 75% to 125% of the average size of the one or more microorganisms.

6. The device according to claim 1 or 2, wherein at least one of the plates is transparent.

7. The device according to claim 1 or 2, further comprising: (a) A housing configured to hold the sample and to be mounted to a mobile communication device.

8. The device of claim 1 or 2, wherein the housing comprises an optical device for facilitating imaging and / or signal processing of the sample by a mobile communication device, and a base configured to hold the optical device on the mobile communication device.

9. The apparatus according to claim 1 or 2, wherein the processor is configured to receive and analyze the series of images, and identify and count the microorganisms in the first region in each of the images.

10. A device according to claim 1 or 2, wherein one or both of the plates comprise a control zone and an experimental zone on the corresponding sample contact area, wherein the experimental zone comprises one or more test reagents that dissolve and diffuse into the sample upon contact with the sample, and wherein the control zone does not comprise such test reagents.

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