Substrates with channels for controlled fluid flow in bioassay sampling - Patent Application 20070122999

Abstract

Immunoassay devices are described that include multiple fluid flow channels that are distinct and designed for optimal fluid control. Substrates configured to control the rate of fluid flow for in situ immunoassay measurements that detect and quantify the presence of one or more analytes of interest in a sample are also described. More specifically, the present disclosure relates to consumables for lateral flow assays that detect markers or causative agents of medical conditions in combination with instruments.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 62 / 959,748, filed January 10, 2020, which is incorporated herein by reference in its entirety.

[0002] Technical Field The subject matter described herein relates to substrates configured to control the rate of fluid flow for in situ immunoassay measurements to detect and quantify one or more analytes of interest in a sample. More specifically, the disclosure relates to consumables for lateral flow assays in combination with instruments to detect markers or causative agents of medical conditions. [Background technology]

[0003] background Lateral flow assays are an established technology that can be adapted for a variety of testing applications for sensors, diagnostics, and indicators. Lateral flow assays typically consist of a material or substrate that transports a fluid sample of interest from an application point (e.g., a sample collection zone) to a detection zone via passive capillary action. For example, rapid lateral flow immunoassay testing devices are used in both clinical and home environments. These devices are used to test for various analytes, such as hormones, proteins, urine, or plasma components. These devices generally include a lateral flow test strip, such as nitrocellulose or filter paper, a sample application area, a test result area, and an analyte-specific binding reagent that is bound to some type of detectable "label" or "reporter," such as a colored particle (e.g., europium beads), a fluorescent or luminescent tag, or an enzyme detection system. The simplicity of these devices contributes to their continued use in the marketplace. Because the method of fluid transport is passive, the predetermined flow path, along with the flow rate, is primarily determined by the viscosity of the liquid sample, the substrate material, and the chemical properties (e.g., hydrophilicity or hydrophobicity) of any coatings that may be applied. It would be advantageous to modify the flow rate or control the uniformity of the fluid flow without adding extra components or materials to the substrate. A technique for modifying and adjusting the flow rate and flow uniformity of a fluid sample deposited on a substrate in a lateral flow assay is desired. [Brief explanation of the drawings]

[0004] BRIEF DESCRIPTION OF THE DRAWINGS [Figure 1]

[0004] An architecture is shown that includes a remote server, a database, and an image capture device that collects images from test strips within a housing, according to some embodiments. [Figure 2A]

[0005] 1 shows an immunoassay device. [Figure 2B]

[0005] Figure 1 shows the progression of a fluid along a device. [Figure 2C]

[0005] Figure 1 shows the progression of a fluid along a device. [Figure 3A]

[0006] 1 shows an immunoassay device according to some embodiments. [Figure 3B]

[0006] Figure 3B shows a substrate of the immunoassay device of Figure 3A with multiple fluid flow channels, according to some embodiments. [Figure 3C]

[0006] Figure 3B shows a substrate of the immunoassay device of Figure 3A with multiple fluid flow channels, according to some embodiments. [Figure 4A]

[0007] 1 shows an immunoassay device according to some embodiments. [Figure 4B]

[0007] Figure 4B shows a substrate of the immunoassay device of Figure 4A with multiple fluid flow channels, according to some embodiments. [Figure 5A]

[0008] 1 illustrates a test strip with multiple fluid flow channels and dimensions of some of its features, according to some embodiments. [Figure 5B] 1 illustrates a test strip with multiple fluid flow channels and dimensions of some of its features, according to some embodiments. [Figure 6A]

[0009] Data is shown showing fluid flow in a test strip with and without a diamond-shaped fluid control feature at the channel inlet, and the change in flow rate depending on whether the user places the fluid sample in the sample zone of the test strip quickly or slowly. [Figure 6B]

[0009] Data is shown showing fluid flow in a test strip with and without a diamond-shaped fluid control mechanism at the channel inlet, and the change in flow rate depending on whether the user places the fluid sample in the sample zone of the test strip quickly or slowly. [Figure 7A]

[0010] 1 illustrates a test strip having a conjugate zone and / or a capture zone comprised of an array of reagent droplets, according to some embodiments. [Figure 7B]1 illustrates a test strip having a conjugate zone and / or a capture zone comprised of an array of reagent droplets, according to some embodiments. [Figure 8A]

[0011] 1 is an image of a test strip with multiple individual, separate fluid flow channels after testing for fluid flow and signal acquisition. [Figure 8B]

[0011] Figure 1 is an image of a test strip with multiple non-discrete fluid flow channels after testing for fluid flow and signal acquisition. [Figure 9A]

[0012] 1 illustrates a test strip with a barrier region according to some embodiments. [Figure 9B] 1 illustrates a test strip without a barrier region, according to some embodiments. [Figure 9C]

[0013] 9C shows results from testing fluid flow in the test strip of FIGS. 9A and 9B, according to some embodiments. [Figure 10A]

[0014] 1 illustrates alternative patterns for creating flow channels and flow in a test strip. [Figure 10B]

[0014] Alternative patterns for creating flow channels and flow in a test strip are shown. [Figure 11]

[0015] 1 is a flowchart illustrating steps of a method for diagnosing, treating, or both, a condition or disorder in a subject, according to some embodiments. [Figure 12]

[0016] 12 is a block diagram illustrating an example computer system capable of implementing the client and server of FIG. 1 and the method of FIG. 11, according to some embodiments. DETAILED DESCRIPTION OF THE INVENTION

[0005] Detailed Description definition

[0017] Various aspects will now be described more fully hereinafter. However, such aspects may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey its scope to those skilled in the art.

[0006]

[0018] When a range of values is provided, each intervening value between the upper and lower limits of that range, and any other stated or intervening value within that stated range, is intended to be encompassed within the disclosure. For example, if a range of 1 μm to 8 μm is stated, then 2 μm, 3 μm, 4 μm, 5 μm, 6 μm, and 7 μm are intended to be expressly disclosed as well, along with ranges of values above 1 μm and below 8 μm.

[0007]

[0019] The singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a polymer" includes a single polymer as well as two or more of the same or different polymers, reference to "an excipient" includes a single excipient as well as two or more of the same or different excipients, etc.

[0008]

[0020] A "sample" is any substance to be tested for the presence or amount of an analyte of interest. Preferably, the sample is a fluid sample, preferably a liquid sample. Examples of liquid samples that can be tested using the test device include bodily fluids including blood, serum, plasma, saliva, urine, ocular fluid, semen, sputum, nasal fluid, and spinal fluid.

[0009] Overall overview

[0021] Embodiments consistent with the present disclosure utilize fabrication techniques to modify nitrocellulose strips into multiple channels with desired fluid flow characteristics. In some embodiments, the nitrocellulose strips provide a substrate for simple yet accurate diagnostic procedures for selected diseases (e.g., Legionella, influenza, Ebola, Lyme disease, etc.). Test types consistent with embodiments in the present disclosure can include any type of spectroscopic analysis of test assays using electromagnetic radiation, such as, but not limited to, absorption spectroscopy (ultraviolet, visible, or infrared), including reflectance or transmittance spectroscopy, or emission spectroscopy, including fluorescence and emission spectroscopy, Raman spectroscopy, and any type of radiation scattering. Furthermore, embodiments as disclosed herein can further utilize the networking capabilities of such appliances to enhance the processing, cataloging, coordination, and cross-referencing capabilities of each test by using cloud computing solutions. Thus, in some embodiments, high-quality (e.g., high spatial and spectral resolution) images, sequences of images, videos, or processed versions thereof are uploaded to a remote server that can perform massively parallel computing to provide diagnostic results in short timeframes. Such analyzed material may be processed immediately or at a later date / time and / or compared to previously collected material to determine differences over time, e.g., the time evolution of analytes across the test strip. After user anonymization, such analyzed material may be used for analysis in the interest of public health or to provide further benefits to users of the test by cross-referencing results with others by predetermined criteria, e.g., age group, sex, geographic location, pathogen characteristics, etc.

[0010]

[0022] The system of the present application offers several advantages, such as allowing the user to quickly know whether a disease is present or potential, or whether it is mild or severe, without the need for a specialist or the use of complex machines or equipment.

[0011]

[0023] While many of the examples provided herein describe downloading and storing a user's identifiable personal information and data, or a user's interaction history with one or more remote clinics, each user may provide explicit permission for such user information to be shared or stored. Explicit permission can be provided using privacy controls integrated into the disclosed system. Each user can be notified that such user information is being shared with their explicit consent, and each user can terminate information sharing and delete any stored user information at any time. Additionally, in some embodiments, stored user information can be encrypted to protect the user's security and identity.

[0012] System architecture example

[0024] FIG. 1 illustrates an architecture 10 including a remote server 110, a database 152, and an image capture device 130 that collects images from a test strip 100 within a housing 135, according to some embodiments. In the architecture 10, the test strip 100 and the housing 135 may be consumable items that a user can dispose of after use. For example, the test strip 100 may be replaced after each use of a test sample, while the housing 135 may be used several more times. In that regard, the test strip 100 and the housing may be part of a package that a user requests from a clinical service provider. The package may include one housing 135 and multiple test strips 100 that can be used therewith. In some embodiments, the housing 135 may be a semi-permanent or premium auxiliary box (e.g., a cassette or cartridge) that can be used multiple times, regardless of whether it is part of a package. In some embodiments, the housing 135 is a housing or cartridge that facilitates handling of the test strip 100. In other embodiments, the test strip 100 is an immunoassay test strip, such as a dipstick. That is, the housing 135 is optional and, if present, can be a flexible laminate such as that disclosed in US Patent Application Publication No. 2009 / 02263854 and shown in Design Patent No. D606664.

[0013]

[0025] In addition to consumables, image capture device 130 may include a smartphone or other mobile computing device (e.g., a tablet, pad, or even a laptop) provided by a user. Image capture device 130 may generally include a sensor array 140 and an optical coupling mechanism 120 (e.g., a lens system with autofocus capabilities). Image capture device 130 may also be configured to wirelessly couple with a remote server 110 and a remote database 152 via network 150. Remote server 110 may provide support for an image capture application 145 installed on image capture device 130. Support may include installation, updates, and maintenance of image capture application 145, acquisition of raw data (e.g., photos, photo sequences, and videos) for storage in database 152, image processing, etc.

[0014]

[0026] Although some of the descriptions herein focus on fluorescence spectroscopy of test strips, some embodiments consistent with the present disclosure may include any other type of electromagnetic interaction and spectroscopic analysis. Some examples of spectroscopic analysis consistent with the present disclosure may include Raman spectroscopy, infrared absorption spectroscopy, infrared reflectance / transmittance spectroscopy, etc. Furthermore, in some embodiments, the emission source may be replaced with a light coupling mechanism (e.g., a lens, a mirror, a prism, a diffraction grating, or any combination thereof) so that solar radiation (e.g., daytime) or any external illumination is used to excite the spectral response of the region of interest of the test strip.

[0015]

[0027] The housing 135 is configured to avoid or control any extraneous light that interferes with the fluorescent excitation light or fluorescent emission light collected by the image capture device. For example, it may be desirable to uniformly illuminate the region of interest on the test strip (e.g., free of shadows, bright spots, or other artifacts) to create a smooth spectral background that can be filtered out by an image capture application in the image capture device.

[0016]

[0028] Some embodiments extract values for assessing the diagnosis of an assay by spatially and / or spectrally filtering an image of the test strip 100. Thus, filtered pixel values can be tallied and compared to a preselected threshold. Thus, if the tallied value is less than or greater than the threshold, a positive disease diagnosis is likely. Some embodiments may include an error value based on statistical analysis and calibration to provide a confidence interval for the diagnosis. In other embodiments, information can be compared between the area occupied by one analyte band and a similar area where there is no uptake of the fluorescent conjugate.

[0017] Substrate containing fluidic channels and fluidic control features

[0029] 2A-2C illustrate the progression of a sample as it is placed on an assay device. The sample may include fluids consisting of a subject sample (e.g., from a patient, test container, etc.) and, if necessary, reagents or processing fluids. The reagents or processing fluids are optional and may be useful to facilitate flow of the sample through different portions of the immunoassay device in a "lateral direction" (e.g., from left to right in the figure) by capillary action. The subject sample may be collected from the patient via a swab (e.g., nasal swab, or other body cavity), syringe, or scoop in a preselected volume, e.g., 100 microliters (μL) or more.

[0018]

[0030] Referring initially to FIG. 1A , immunoassay device 200 includes substrate 202. Deposited or formed on the substrate are a sample zone 204 in fluid communication with a conjugate zone 206, a capture zone 208, and an optional absorbent pad 210. Typically, conjugate zone 206 is downstream of sample pad 204, capture zone 208 is downstream of conjugate zone 206, and absorbent pad 210 is downstream of capture zone 208. In some embodiments, an image capture device is configured to capture and process an image of at least a portion of capture zone 208 (e.g., image capture device 130; see FIG. 1 ). Accordingly, the light source can be configured to excite a signal, such as fluorescence, from the test strip. In some embodiments, the emitted signal, such as fluorescence, has a wavelength within a color range selected for a sensor array in the image capture device.

[0019]

[0031] In some embodiments, the conjugate zone 206 comprises a mobile detectable species. Examples of mobile detectable species are known in the art and depend on the analyte of interest (e.g., an infectious agent, or a chemical moiety such as a drug or pollutant). In some embodiments, the immunoassay device does not have a conjugate zone, and the mobile detectable species is provided in a container with the immunoassay device, for example, as a lyophilized material. The sample and lyophilized material are mixed, and the mixture is deposited on the sample pad 204.

[0020]

[0032] In some embodiments, the capture zone 208 includes one or more lines, bands, or spots, such as the first control zone 212, the first test zone 214, and the second test zone 216 (collectively referred to herein as "capture zones"). Thus, the shape and number of capture zones can include multiple variations, i.e., dots, droplets, lines, and arrays of dots and / or lines, as well as curved shapes with more complex shape factors. The capture zones contain at least one immobilizable species that has a chemical or physical affinity for at least a portion of the conjugate complex formed between the mobile detectable species and the analyte of interest or a control analyte. The binding species in each capture zone are deposited or printed from solution and allowed to dry for a period of time (e.g., minutes, hours, or overnight). In some embodiments, each control or test line within a capture zone contains a binding member for a specific analyte, with each analyte binding to a distinct mobile detectable species, and the detectable species differ in signal emission, e.g., wavelength or type. In one embodiment, each control or test line binds a conjugate of the analyte and a mobile detectable species that generates a light signal at a different wavelength. Exemplary immunoassay test strips are described, for example, in U.S. Patent Nos. 9,207,181, 9,989,466, and 10,168,329, and U.S. Patent Application Publication Nos. 2017 / 0059566 and 2018 / 0229232, each of which is incorporated herein by reference.

[0021]

[0033] The immunoassay device 200 can be uniquely configured for the detection of specific pathogens or analytes of a species of interest. These pathogens or analytes include, but are not limited to, proteins, haptens, immunoglobulins, enzymes, hormones, polynucleotides, steroids, lipoproteins, drugs, bacterial antigens, and viral antigens. With respect to bacterial and viral antigens, more commonly referred to in the art as infectious antigens, analytes of interest include streptococci, influenza A, influenza B, respiratory syncytial virus (RSV), hepatitis A, hepatitis B, and / or hepatitis C, pneumococcus, human metapneumovirus, and other infectious agents known to those skilled in the art. In some embodiments, the test device is intended to detect one or more antigens associated with Lyme disease. In some embodiments, the immunoassay device is intended for use in the field of women's health. For example, test devices are contemplated that detect one or more of fetal fibronectin, chlamydia, human chorionic gonadotropin (HCG), hyperglycosylated chorionic gonadotropin, human papillomavirus (HPV), etc. In another embodiment, the immunoassay device is configured to detect vitamin D and is designed to interact with the normalization apparatus and methods described herein. Techniques used to measure signals from the immunoassay device can include any immunoassay technique, such as non-competitive assay techniques, competitive assay techniques (e.g., homogeneous competitive assays, heterogeneous competitive assays), etc.

[0022]

[0034] In some embodiments, the analytes of interest may include disease-transmitting pathogens such as respiratory syncytial virus (RSV), influenza A virus, influenza B virus, or human metapneumovirus (hMPV). In some embodiments, the analytes of interest may include controlled substances such as drugs and other illegal or prohibited substances (e.g., steroids, etc.). For example, some embodiments may include the detection and measurement of drugs such as fentanyl, buprenorphine, oxycodone, and / or 7-aminoclonazepam.

[0023]

[0035] Continuing with FIG. 2A , sample pad 204 receives a sample suspected of containing an analyte of interest. Conjugate zone 206, in some embodiments, contains two dried conjugates, each containing a particle comprising a detectable label element, such as a fluorescent element. Exemplary fluorescent elements are lanthanides, such as one of the following 15 elements: lanthanum, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, terbium, dysprosium, holmium, erbium, ytterbium, lutetium, and yttrium. In one embodiment, the lanthanide is embedded within or on a particle, such as a polystyrene particle. The particle can be a microparticle (a particle less than about 1,000 micrometers in diameter, in some cases less than about 500 micrometers in diameter, and in some cases less than 200, 150, or 100 micrometers in diameter) comprising a luminescent or fluorescent lanthanide; in some embodiments, the lanthanide is europium. In some embodiments, the lanthanide is chelated europium. The microparticles have a lanthanide core with a polymer coating, such as a europium core with a polystyrene coating, in some embodiments. A binding partner for the analyte of interest in the sample is attached to or associated with the outer surface of the microparticle. In some embodiments, the binding partner for the analyte of interest is an antibody, monoclonal antibody, or polyclonal antibody. Those skilled in the art will recognize that other binding partners can be selected, and that such binding partners can include conjugates, such as biotin and streptavidin conjugates. Upon entering the conjugate zone 206, the liquid sample hydrates, suspends, and mobilizes the dried microparticle-antibody conjugates, transporting the conjugates, along with the sample, downstream on the test strip to the control or reference and / or test lines in the capture zone 208.As the sample and microparticle-antibody conjugates continue to flow downstream on the test strip, if the analyte of interest is present in the sample, the fluorescent microparticle-antibody conjugates, now bound to the antigen / analyte of interest, will bind to the predetermined binding member for the analyte of interest immobilized on the conjugate line in conjugate zone 208. In some embodiments, a single test line is present on the immunoassay substrate. In some embodiments, at least two or more test lines are present. By way of example, capture zone 208 may be designed to detect and / or differentiate between influenza A and influenza B and may include a first test line 214 for detecting influenza A and a second test line 216 for detecting influenza B. Microparticle-antibody conjugates, including microparticles coated with antibodies specific for influenza A and microparticles coated with antibodies specific for influenza B, may be included in conjugate zone 206, in some embodiments downstream of control line 212. The first test line for influenza A contains a monoclonal or polyclonal antibody directed against a determinant on the nucleoprotein of influenza A, and the second test line for influenza B contains a monoclonal or polyclonal antibody directed against a determinant on the nucleoprotein of influenza B. If antigen is present in the sample, a typical immunoassay sandwich is formed with each test line corresponding to an antigen in the sample.

[0024]

[0036] Immunoassay test devices are intended to receive a wide variety of samples, including biological samples from human bodily fluids, including, but not limited to, nasal secretions, nasopharyngeal secretions, saliva, mucus, urine, vaginal secretions, fecal samples, blood, etc. In some embodiments, the kits described herein are provided with a positive control swab or sample. In some embodiments, a negative control swab or sample is provided. For assays requiring external positive and / or negative controls, the user may be prompted to insert or apply a positive or negative control sample or swab.

[0025]

[0037] When an immunoassay band is immobilized on a substrate by attachment (e.g., adsorption, chemisorption, immunoligand, etc.) to an immunoprotein in the immunoassay strip, it fluoresces primarily from a fluorescent dye bound to the target analyte. Therefore, any red emission within the band boundary is largely due to the presence of the target analyte (e.g., the presence of a pathogenic antigen, etc.). However, the amount of red signal within the immunoassay band boundary may include some background. To better assess background signal (e.g., not caused by target analyte bound to antibodies on the band), some test strips may include a blank control area.

[0026]

[0038] In some embodiments, the first and second control lines are positioned on either side of the test line relative to the flow direction. Thus, the first and second control lines provide start / stop signals for the assay. This allows for a negative result to be predicted even if only a portion of the second control line is wetted with sample. In some embodiments, the image capture device can capture a pixelated image of the capture zone 208, thus capturing the progress of the fluid front over time as it flows downstream from the sample zone across the test substrate. Thus, some embodiments can provide metrics and performance data for the substrate as the progress of the sample fluid is tracked. This concept is described with reference to FIGS. 2B-2C, which show a moving fluid front 218. The time it takes for the fluid front to travel between two points on the test strip can be determined, for example, by analyzing image frames collected during the flow of sample between the two points. In addition to determining the fluid flow rate, the shape (e.g., curvature and slope) of the fluid front 210 can be visualized in the image as a metric for the test strip.

[0027]

[0039] The test device shown in FIG. 2B illustrates a sample deposited on the sample pad 204 and flowing across the conjugate pad 206 into the capture zone 208. In the device of FIG. 2C, the sample is progressing through the capture zone 208. In some embodiments, it is desirable to perform measurements on the sample after the fluid front 210 reaches a landmark location. Thus, in some embodiments, the complete progression of the fluid front 210 along the substrate can be recorded by an image capture device. In some embodiments, it may be desirable for the fluid front 210 to form a line substantially perpendicular to the progression of the sample to ensure that the sample contacts substantially the entire width of the test line in the capture zone approximately simultaneously. In some embodiments, the fluid front 210 can have any shape (concave, convex, irregular, etc.), and the image capture device can be configured to follow the fluid front 210 as it progresses across the capture zone. The time for the sample to flow from the sample zone to the end of the capture zone can be 5 minutes, 7 minutes, 10 minutes, 12 minutes, 15 minutes, 17 minutes, 20 minutes, or more.

[0028]

[0040] The substrate of the immunoassay device may be a laminate consisting of a first base or support layer and a second membrane layer, where the first support layer can be hydrophobic or hydrophilic, and the second membrane layer is adhered to the first layer, the second membrane layer being hygroscopic in nature and / or capable of capillary flow. The support layer may be hydrophobic or impermeable, such as polyethylene terephthalate, polyester, silicone, etc.

[0029]

[0041] With the introduction provided in Figures 2A-2C of a schematic immunoassay device, the immunoassay device of the present disclosure will now be described. In a first embodiment, the immunoassay device is comprised of a single, integral substrate containing multiple separate fluid flow channels. The substrate may be a laminate material, as described above, and is now a single, integral laminate substrate configured as described with reference to Figures 3A-3C. Figure 3A shows an immunoassay device 300 comprised of a test strip 302 inserted into an optional housing 304. Figure 3B shows test strip 302. Test strip 302 includes a single sample zone 304 accessible via port 306 within the optional housing of Figure 3A. Single sample zone 304 is common to and in fluid communication with multiple fluid flow channels, such as the individual, separate channels identified as 308, 310, 312, and 314. Each fluid flow channel is in direct or indirect fluid communication with the sample zone at a channel inlet region of each fluid flow channel, such as channel inlet 316 of channel 308 .

[0030]

[0042] Each fluid flow channel in the plurality of channels has a length l fc and width w fc Each fluid flow channel includes a capture zone downstream from the channel inlet region and a channel constriction zone positioned between the channel inlet region and the capture zone, the channel constriction zone having a width w cz and length l cz and the width of the channel constriction zone w cz is a minimum value equal to or greater than (i) the diameter of the particle reagent deposited or to be deposited on the substrate, or (i') the width w of the fluid flow channel fc and (ii) a width of the fluid flow channel, w fc This corresponds to a value within a range determined by a maximum value less than or equal to about 75% of the value.

[0031]

[0043] 3B and 3C, the fluid flow channel 308 has a width wfc and length l fc The fluid flow channels also include a capture zone 318 positioned downstream from the channel entrance region 316 and upstream from a channel constriction zone, shown generally across the plurality of fluid flow channels in FIG. 3B as 320 and shown in FIG. 3C as constriction 322 of fluid flow channel 308. A constriction in each fluid flow channel, such as constriction 322, has a width w cz and length l cz In embodiments where the width of the constriction varies along its length, the width w cz is the length l cz 3C , downstream of the channel constriction zone is a capture zone 323 containing one or more capture or test or control "lines." The use of the term "line" is not intended to convey the geometric shape of the line, as the lines can be any geometric shape, such as circular, diamond, triangular, or an array of any geometric shape. In the fluid flow channel 308 of FIG. 3C , downstream of the constriction 322 are three capture lines 324, 326, 328.

[0032]

[0044] Test strip 302 in FIGS. 3B and 3C is a monolithic substrate, i.e., the substrate is a single, continuous material. In one embodiment, the single, continuous substrate is a laminate of a support material and a membrane material. The membrane material is processed to form a fluid flow channel including a constriction region and a capture line, as described herein. In one embodiment, the membrane material is a bibulous material, such as nitrocellulose. The nitrocellulose is exposed to a chemical or laser to remove or etch portions of the nitrocellulose to form the flow channel. For example, fluid flow channel 308 in FIGS. 3B and 3C is formed by etching away membrane material to form opposing sidewalls 330, 332. The dimensions of the sidewalls are tailored at the constriction region, as can be seen. The test strip can further include a fiducial, such as fiducial 334, and a lateral barrier, such as barrier 336. The fiducial aids in optical analysis of the conjugate line, and the barrier aids in controlling fluid flow through the test strip. In one specific embodiment, a laser is used to ablate substrate film material in a controlled manner. Laser ablation generally refers to the process of removing material using incident light of a certain wavelength. For example, in polymeric materials, the incident light generally induces photochemical changes in the polymer that result in chemical dissolution. Any known laser can be employed in the present invention, including, for example, CO2 lasers, pulsed light lasers, diode lasers, ND:Yag 1064 nm and 532 nm lasers, alexandrite and Q-switched lasers, pulsed dye lasers, optical and RF lasers, erbium lasers, ruby lasers, and holmium lasers. In a preferred embodiment, a CO2 laser is used to etch a nitrocellulose film attached to a support fixture. Using a moving beam or XY table, precise channels are created in the nitrocellulose to define, for example, fluid flow channels or other fluid features. Additionally, other optical devices, such as optical lenses and mirrors, may be employed in combination with the laser to enhance channel formation.In another embodiment, a Nd:YVO solid state laser with picosecond pulses is used, for example, at a wavelength of 532 nanometers, a pulse length of 12 picoseconds, a pulse energy of 10 microjoules, and a pulse frequency of 10 kilohertz, using a 100 millimeter F-theta lens and a feed rate of 25 milliseconds per second to focus the beam onto the substrate 301. The parameters for laser ablation of a substrate, such as wavelength, pulse duration, pulse repetition rate, and beam quality for any given laser, can be determined by one skilled in the art.

[0033]

[0045] 3C, each fluid flow channel is physically separated from an adjacent fluid flow channel by a gap g, which corresponds to the area of the substrate that has been ablated or to the thickness / width of the sidewall. In some embodiments, g may have a dimension of at least about 0.01 mm, 0.025 mm, 0.03 mm, 0.05 mm, 0.07 mm, 0.08 mm, 0.09 mm, 0.1 mm, 0.2 mm, 0.3 mm, 0.4 mm, 0.5 mm, 0.6 mm, 0.7 mm, 0.8 mm, 0.9 mm, or 1 mm, or between about any two of these discrete values.

[0034]

[0046] In some embodiments, the width w of each fluid channel fc may have a dimension of about 0.1 mm, 0.2 mm, 0.3 mm, 0.4 mm, 0.5 mm, 0.6 mm, 0.7 mm, 0.8 mm, 0.9 mm, 1 mm, 1.1 mm, 1.2 mm, 1.3 mm, 1.4 mm, 1.5 mm, 1.6 mm, 1.7 mm, 1.8 mm, 1.9 mm, or 2 mm, or between about any two of these discrete values. For example, in some embodiments, w fc may be 1 mm and w2 may be 0.5 mm.

[0035]

[0047] In some embodiments, the capture lines or dots are deposited on the substrate using inkjet techniques, such as those used in the printing industry. As discussed above, in some embodiments, the capture lines can include an array of dots, where the array has dimensions of m×n (e.g., columns×rows). Thus, in some embodiments, the capture zones on the test strip and / or capture lines in individual fluid flow channels can include an m×n array of separate droplets or dots, where m and n are 1 or greater and each dot in the m×n array is separated from an adjacent dot by a distance x (e.g., "pitch" or "spacing"). In some embodiments, m and n can be any integer, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more. In some embodiments, the pitch x can be about 20-1000 μm, or about 50-500 μm, or about 75-500 μm, or about 100-500 μm, or about 150-500 μm, or about 150-300 μm, or about 150-250 μm, or about 200-500 μm. In some embodiments, the volume of the formulation deposited onto the substrate to form each dot 325 can be about 20-1000 pL, or between about 50-800 pL, or between about 75-800 pL, or between about 100-600 pL, or between about 150-550 pL, or between about 200-500 pL, or between about 200-450 pL.

[0036]

[0048] In some embodiments, each dot in the array can contain a different immobilizable detectable species, arranged in an m×n array to optimize capture / detection efficiency and also to provide and improve measurement quantification. For example, in some embodiments, dots along the same fluidic channel 308 (e.g., along the same column in an m×n array) can contain the same immobilizable detectable species. Thus, as sample fluid progresses through the columns of dots in the fluidic flow channel, the attenuation of the signal collected from the dots along each fluidic channel can be mathematically fitted to a model according to the exact value of the analyte concentration in the sample (given the known concentration of the detectable species on the dots). In some embodiments, dots along the same column can contain the same immobilizable detectable species, while each column is associated with a different species. Thus, multiple analytes of interest can be detected along each of the fluidic flow channels, and multiple flow channels allow for statistical comparison of measurements.

[0037]

[0049] In some embodiments, conjugates specific to each assay (e.g., RSV, Flu A, Flu B, hMPV, etc.) are printed separately onto the capture line of the conjugate zone of each fluid flow channel, thus providing a test assay for multi-analyte detection. Each drop of the array is deposited onto the substrate from a precision liquid dispensing instrument. In some embodiments, the dispensing instrument allows the user to select the droplet volume, droplet pitch, and other variables. The user can also select whether multiple droplets are deposited at each location in the array in a single pass of the instrument dispensing head or in multiple passes of the instrument dispensing head. Different instrument variables can be adjusted to achieve the desired positional accuracy of each dot in the array. Some of the instrument variables can include selecting the number of droplets of formulation containing a conjugate to a mobile detectable species dispensed to form each droplet. In some embodiments, each droplet of the array can be formed by dispensing 1, 5, 10, or more droplets of the formulation. The volume of the droplets can be adjusted as desired, for example, by tens or hundreds of picoliters (pL) per droplet. Furthermore, in some embodiments, multiple dots in the array can be deposited in several passes of the jetting head. For example, a first number of droplets can be deposited on each dot in a first pass, and a second number of droplets can be deposited on each dot in a second pass. In some embodiments, depositing fewer droplets in multiple passes of the dispensing head can improve the placement accuracy of the droplets in the array and result in a more uniform pitch (horizontal and vertical).

[0038]

[0050] FIGS. 4A and 4B show another exemplary immunoassay test device 400. The immunoassay device 400 includes an optional housing 402 for a test strip 404, which is shown separated from the housing in FIG. 4B. The test strip 404 includes a single, integrated substrate 406. As described above, the substrate 406 can be a laminated material consisting of a support layer and a bibulous layer. The bibulous layer is treated or fabricated to include multiple separate fluid flow channels. The embodiment shown in FIGS. 4A and 4B includes four separate fluid flow channels, with channel 408 being representative. The substrate also includes a single sample zone or region 410 in direct fluid communication with each of the multiple channels. More specifically, the channel inlet region of each channel, such as channel inlet region 412 of representative channel 408, is in direct fluid communication with the sample zone that receives the sample for analysis, without any intervening structures, materials, zones, and / or components. Each fluid flow channel in the plurality of channels includes a conjugate zone, shown at 414 in representative channel 408. Each fluid flow channel in the plurality of flow channels includes a capture zone, shown at 416 in representative channel 408. The capture zone includes one or more test lines, such as line 418, that contain immobilized reagents, as described above.

[0039]

[0051] Each fluid flow channel in the plurality of channels also includes a fluid control zone 420, which may be referred to as a constriction zone in embodiments where the fluid control zone is designed to slow or restrict fluid flow. The fluid control zone 420 is positioned between the channel inlet and the conjugate zone. In the embodiment of Figure 4B, the fluid control zone is downstream of the conjugate zone, but it may also be upstream of the conjugate zone.

[0040]

[0052] Test strip 404 is also configured to include features for directing and / or metering a fluid sample disposed on the sample zone into the fluid flow channels. These features are fluid barriers 422, 424. The dimensions of each barrier and the position of each barrier can be adjusted to direct and / or meter fluid into each fluid flow channel. The width of the barrier, shown as w, and its angle α can be varied and selected to adjust fluid dynamics. In one embodiment, the barriers are dimensioned to direct and meter a volume of fluid sample disposed on the sample zone into each fluid flow channel to achieve a substantially uniform flow rate in each of the multiple channels (e.g., a flow rate that varies across the multiple channels by less than about 15%, 10%, or 5%) and / or containment of the volume of fluid sample in the sample zone and fluid flow channels (and ultimately in any absorbent zones at the ends of the channels). That is, none of the fluid sample overflows, seeps, or spills into the area identified by 426 in FIG. 4B.

[0041]

[0053] The channel inlet region of each individual fluid flow channel can be sized and configured to regulate and control the entry of a portion of the fluid sample into each channel. In some embodiments, the channel inlet region includes a fluid control feature, such as a constriction zone 430. The fluid control feature can have any desired shape, such as a quadrilateral (e.g., rhomboid), hourglass, or diamond shape.

[0042]

[0054] The fluid control zones, or channel constriction zones if designed to slow fluid flow, are dimensioned to achieve fluid control in each fluid flow channel, as discussed now with reference to Figures 5A and 5B. Test strip 500 is shown, which is constructed from a unitary, continuous piece of material, which is processed as described herein to define multiple fluid flow channels. Each fluid flow channel has a length l c (Fig. 5B) and width w c(FIG. 5A). The channel constriction zone in each fluid flow channel has a length lcz (FIG. 5B) and a width w cz (FIG. 5A). In one embodiment, the width w cz is a value within a range having a minimum and a maximum value. In one embodiment, the minimum value of the range is equal to or greater than the diameter of the particle reagent deposited on the substrate or to be deposited on the substrate. For example, as described above, the conjugate zone in each fluid flow channel can contain mobile detectable particles. The width w of the constriction zone czis a dimension that allows the mobile, detectable particles to flow through the constriction zone. For example, polymer particles bearing a detectable label used as reagents in the device have an outer diameter. In embodiments in which the immunoassay device is configured such that the conjugate zone is upstream of a constriction zone containing mobile, optically detectable solid particles of a certain diameter, or where the immunoassay device is designed to contain a reagent of a certain dimension that must flow through the constriction zone, the minimum width of the constriction zone is equal to or greater than the dimension of the reagent that must flow therethrough. Exemplary minimum widths of the constriction zone are about 0.01 to 750 micrometers (0.00001 to 0.75 mm), 0.01 to 500 micrometers, 0.01 to 250 micrometers, 0.01 to 100 micrometers, 0.01 to 50 micrometers, 0.01 to 25 micrometers, 0.01 to 10 micrometers, 0.01 to 5 micrometers, 0.01 to 2 micrometers, 0.01 to 1.5 micrometers, 0.01 to 1.0 micrometers, 0.05 to 750 micrometers, 0.05 to 500 micrometers, 0.05 to 250 micrometers, 0.05 to 1.0 micrometers, 0.05 to 100 micrometers, 0.05 The range is 0.075 to 50 micrometers, 0.05 to 25 micrometers, 0.05 to 10 micrometers, 0.05 to 5 micrometers, 0.05 to 2 micrometers, 0.05 to 1.5 micrometers, 0.05 to 1.0 micrometers, 0.075 to 500 micrometers, 0.075 micrometers, 0.075 to 250 micrometers, 0.075 to 100 micrometers, 0.075 to 50 micrometers, 0.075 to 25 micrometers, 0.075 to 10 micrometers, 0.075 to 5 micrometers, 0.075 to 2 micrometers, 0.075 to 1.5 micrometers, or 0.075 to 1.0 micrometers.

[0043]

[0055] Alternatively, the minimum value of the dimensional range of the fluid flow channel width in the constriction zone may be the fluid flow channel width w cFIG. 5A shows a cross section of the fluid flow channel having a width w c and the flow channel width in the constriction zone w cz According to this embodiment, w cz Ha w c In another embodiment, w cz Ha w c is 0.10 times or more, and w cz Ha w c is 0.15 times or more, and w cz Ha w c is 0.20 times or more, and w cz Ha w c is 0.30 times or more, and w cz Ha w c is 0.35 times or more, and w cz Ha w c is 0.40 times or more, and w cz Ha w c In other embodiments, w cz Ha w c is equal to approximately 0.05 to 0.75 times w cz Ha w c is equal to approximately 0.1 to 0.75 times w cz Ha w c is equal to approximately 0.1 to 0.5 times w cz Ha w c is equal to approximately 0.15 to 0.5 times w cz Ha w c is equal to approximately 0.15 to 0.5 times w cz Ha w c is equal to approximately 0.15 to 0.4 times w cz Ha w c is equal to approximately 0.2 to 0.75 times w cz Ha w c Equal to approximately 0.2 to 0.5 times w cz Ha w c is approximately 0.2 to 0.4 times the

[0044]

[0056] As mentioned above, the width w czis a value within a range having a minimum value and a maximum value. In one embodiment, the maximum value is equal to or less than about 75% of the fluid flow channel width. In other embodiments, the maximum value is equal to or less than about 85%, 80%, 70<65%, 55%, 50%, 45%, 40%, 35%, 30%, 25% of the fluid flow channel width.

[0045]

[0057] As an example, consider a fluid flow channel with a length of 17.70 mm and a width of 1.10 mm. If the assay uses a reagent that is a solid of a size that must flow through the constriction zone, e.g., a solid, optically detectable particle having a diameter of about 0.05-10 micrometers, then the minimum width of the channel at the constriction zone corresponds to the diameter of the particle. Alternatively, if the assay does not have a solid reagent that must flow through the constriction zone, then the minimum width of the channel at the constriction zone is equal to or greater than about 25% of the fluid flow channel width, or in this hypothetical example, 25% of 1.10 mm, which is 0.28 mm or greater. The maximum width of the channel at the constriction zone, width w cz is less than or equal to about 75% of the fluid flow channel width, or 75% of 1.10 mm, which is less than or equal to 0.825 mm. Thus, the minimum and maximum range for the channel width in the constriction zone of this hypothetical channel is 0.28 mm to 0.825 mm, inclusive.

[0046]

[0058] The channel length l c ( FIG. 5B ) in the constriction zone, in one embodiment, corresponds to a value within a range determined by (i) a minimum value equal to or greater than about 8% of the fluid flow channel length and (ii) a maximum value equal to or less than about 75% of the fluid flow channel length. Returning, by way of example, to the hypothetical fluid flow channel described in the preceding paragraph, having a fluid flow channel length (l c ) of 17.70 mm and a fluid flow channel width (w c ) of 1.10 mm, the channel length (l c ) in the constriction zone would be 1.4 mm, at least about 8% of 17.70 mm, and 13.28 mm, no greater than about 75% of 17.70 mm. Thus, the minimum and maximum ranges for the channel length in the constriction zone of this hypothetical channel are 1.4 mm to 13.28 mm, inclusive. In other embodiments, the minimum value within the range for the length of the flow channel in the constriction zone, l c , is equal to or greater than about 2%, 3%, 4%, 5%, 7.5%, 9%, 10%, 12%, 15%, 18%, 20%, 22%, 25%, 28%, 30%, 32%, 35%, 40%, 45%, or 50% of the fluid flow channel length. In other embodiments, the maximum value within the range for the length of the channel in the constriction zone, l c , is equal to or less than about 95%, 90%, 85%, 80%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, or 30% of the fluid flow channel length.

[0047]

[0059] The dimensions and configuration of the channel in the constriction zone can vary along the channel length l c z in the constriction zone. In some embodiments, the entrance and / or exit regions of the constriction zone are shaped to have a taper. The taper can be curved or angled. In one embodiment, the taper extends from the fluid flow channel width to the channel constriction zone width.

[0048]

[0060] In some embodiments, the channel inlet region of each fluid flow channel is configured with a fluid control feature. For example, referring to FIG. 4B, the channel inlet region 412 of representative channel 408 can optionally include a fluid control feature. For example, in test strip 500 of FIGS. 5A and 5B, a common shared sample zone 502 is in direct fluid communication with an inlet region to each of the multiple fluid flow channels, such as representative inlet region 504 to representative channel 506 (FIG. 5A). Inlet region 504 in this embodiment is a constricted region resulting from the diamond-shaped, angled sidewalls of the fluid flow channel in this region. The inlet constriction region has a width w1 (FIG. 5A) and a length L1 (FIG. 5B), and in one embodiment, the inlet constriction region width w1 is greater than the width w of the channel constriction zone. cz In one embodiment, the entrance constriction region width w1 is essentially the same as the width w of the channel constriction zone. cz The width of the entrance constriction region w1 is between the minimum and maximum values set forth above. In one embodiment, the entrance constriction region width w1 has a geometric shape that can be angled or non-angled. In one embodiment, the entrance constriction region includes an angle that is about 30 to 90 degrees. In other embodiments, the entrance constriction region has a geometric shape that is a half diamond, half rectangle, half square, quarter square, quarter rectangle, half parallelogram, quarter parallelogram, or half kite.

[0049]

[0061] As discussed above with respect to FIG. 4B , barrier regions (also referred to as barrier extension regions) are formed in the substrate to direct and control fluid disposed in the sample zone into each of the plurality of fluid flow channels. FIG. 5B provides additional details regarding the barrier regions, consistent with some embodiments. In test strip 500 of FIG. 5B , each fluid flow channel n (such as representative fluid flow channel 506) has a length lc. Each fluid flow channel has opposing sidewalls, such as sidewalls 508, 510 of representative flow channel 506 of FIG. 5B . Each sidewall has a width w s (FIG. 5B). In one embodiment, the barrier region has a thickness of about w s ~about 10w s , or about w s ~About 5ws The width w b It has.

[0050]

[0062] The number of channels of the plurality of fluid flow channels in the test assay can be in the range of 1 to 100, 1 to 50, 1 to 25, 1 to 20, 1 to 15, 2 to 100, 2 to 50, 2 to 25, 2 to 20, 2 to 15, 2 to 10, 3 to 100, 3 to 50, 3 to 25, 3 to 15, or 3 to 10 (including any integer therein), which includes, for example, but not limited to, at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 individual, separate fluid flow channels.

[0051]

[0063] Test strips were prepared and tested as described herein to evaluate fluid flow rate and uniformity across multiple fluid flow channels. In one study, test strips were prepared essentially as depicted in Figures 5A and 5B, with a diamond-shaped constriction feature in the channel inlet region. Test strips without a diamond-shaped constriction feature were also prepared. A known volume of fluid was deposited onto a common sample zone either immediately in one rapid deposition from a pipette or slowly over approximately 15 seconds. The speed of sample deposition onto the sample zone was varied to mimic how different users might deposit samples onto a test device. Figures 6A and 6B show the standard deviation (Figure 6A) and coefficient of variation (Figure 6B) from analysis of fluid flow through test strips with and without a diamond-shaped fluid control feature in the fluid flow channel inlet region. Figure 6A shows the standard deviation of the net signal at fast and slow sample addition rates for test strips with and without a diamond-shaped constriction feature. 6A shows the coefficient of variation of net signal for fast and slow sample addition rates for test strips with and without a diamond-shaped constriction zone. Each data point in the chart corresponds to multiple measurements (e.g., 20 or more) using an equal number of similarly designed test strips. It can be seen that the embodiment with the entrance constriction exhibits reduced strip-to-strip variation compared to the embodiment without the entrance constriction.

[0052]

[0064] 7A and 7B show conjugate zones in a fluid flow channel, where the conjugate zone is an array of dots. Conjugate zone 700 in FIG. 7A is comprised of a 1×8 array of reagent droplets containing a mobile detectable species. Conjugate zone 702 in FIG. 7B is comprised of a 3×10 array of reagent droplets containing a mobile detectable species. It will be understood that each dot need not be of the same composition, and that the capture zone of a fluid flow channel with an immobilizable species can similarly be an array of reagent droplets. The reagent compositions deposited to form the capture zone array or conjugate zone array may include a mobile detectable species, or may include binding partners or species immobilized on a substrate, or may include species useful as controls. The array, in one embodiment, comprises m droplets in one direction and n droplets in a second direction, forming an m×n array, where m and / or n are 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30. In one embodiment, n and m are the same, and in another embodiment, n and m are different values.

[0053]

[0065] In other embodiments, the volume of the formulation deposited on the substrate to form each drop 724 can be about 20-1000 pL, or about 50-800 pL, or about 75-800 pL, or about 100-600 pL, or about 150-550 pL, or about 200-500 pL, or about 200-450 pL.

[0054]

[0066] 8A and 8B are images of a test strip with multiple individual, separate fluid flow channels (FIG. 8A) and a test strip with multiple, non-separate fluid flow channels (FIG. 8B) after testing for fluid flow. FIG. 8A shows an image of test strip 800a with separate individual fluid flow channels 805a-1, 805a-2, 805a-3, and 805a-4 (collectively "fluid flow channels 805a"). FIG. 8B shows an image of test strip 800b without separate fluid flow channels; i.e., the fluid flow channels are not isolated or separated from one another along the length of the channels from their inlet in the common sample zone to their outlet in the common outlet region. Each of the test strips has conjugate zones 821a and 821b, respectively. Hereinafter, test strips 800a and 800b are collectively referred to as "test strip 800." Test strip 800 includes a barrier 827. Test strip 800a includes a constriction region 820. Differences in performance can be seen in the capture regions 823a, 823b of each strip. The test strip of FIG. 8A, with its individual, separate channels and constriction region, provides bright, well-resolved signal brightness in capture zone droplets 825a-1, 825a-2, 825a-3, and 825a-4 (collectively referred to as "droplets 825a"). The signal produced by test strip 800b is less bright and has poorer resolution than that of test strip 800a. Furthermore, non-uniformity in fluid flow is evident from streaks 805b-1, 805b-2, 805b-3, and 805b-4 (collectively referred to as "streaks 805b"). Thus, in some embodiments, the individual, separate fluid channels in the multiple channels are a contributing mechanism for higher, better-resolved signals due to flow uniformity.

[0055]

[0067] 9A and 9B show test strip 900a with barrier 927 and test strip 900b without barrier 927. Test strips 900a and 900b are collectively referred to as "test strip 900." Test strip 900a includes fluidic channels 905-1a, 905-2a, 905-3a, and 905-4a (collectively referred to as "fluidic channel 905a"). Similarly, test strip 900b includes fluidic channels 905-1b, 905-2b, 905-3b, and 905-4b (collectively referred to as "fluidic channel 905b"). Performance of these test strips was compared by depositing a fluid sample in the sample zone and observing the signal in the capture zone. FIG. 9C shows the results of an investigation into test strip 900. The results for test strip 900a are shown in the left panel (901a) of Figure 9C, and the results for test strip 900b are shown in the right panel (901b). Curves 951a and 952a show the signal and background measurements, respectively, for each of channels 905a. Curves 951b and 952b show the signal and background measurements, respectively, for each of channels 905b.

[0056]

[0068] Removal of barrier 927 allows fluid sample deposited in the sample zone to flow toward absorbent pad 912 of test strip 900b. The initial raw signal intensities (curves 951a and 952a) show higher signal and background in the outer channels of test strip 900a. Net signal 961a (obtained by subtracting curve 952a from curve 951a) results in lower net signal in the outer channels (e.g., channels 905-1a and 905-4a). In some embodiments, sample-dependent bias can also be observed. This may include a higher background signal (e.g., curve 952a) with a higher analyte concentration in the sample stream. Thus, test strip 900b without barrier 927 may include significant compensation for channel bias (e.g., curves 951b, 952b, and 961b are straighter and more horizontal).

[0057]

[0069] Another study was performed on test strips with and without barriers. Test strips with four individual, separate fluid flow channels were prepared, one with a barrier and one without. After flowing a fluid containing a detectable species through the sample zone, the signal from the capture zone was evaluated. The barriers were found to improve the uniformity of fluid flow across the channels (data not shown).

[0058]

[0070] As described above, the substrate of the test strip can be a laminate of a support member and a bibulous membrane. Also as described above, the bibulous membrane can be fabricated or treated to etch away portions of the membrane, creating multiple, separate, individual fluid flow channels and fluid control features. Once the membrane is etched away, fluid moving through the membrane contacts the support member. The hydrophobicity and hydrophilicity of the support member can be selected and optimized for fluid control. Additionally, the design of the fluid channel and fluid control features can be varied to control the rate of fluid flow. Some variations are shown in Figures 10A and 10B. Figure 10A shows a test strip 1100 having a substrate 1101 with a serpentine fluid channel 1105 according to some embodiments. The fluid channel 1105 is a flow-reducing structure designed to induce more sample material to interact with the target capture zone 1125-2, thereby improving the detection sensitivity of low-positive results from the assay. By adding a first control capture zone 1125-1 near the sample pad 1111 at the start of the assay and a second control capture zone 1125-3 near the absorbent pad 1112 at the end of the assay, precise start and end points for the test are provided.

[0059]

[0071] 10B shows a test strip 1200 including a mixing longitudinal channel 1205-1 and a serpentine channel 1205-s (collectively referred to herein as "fluidic channels 1205") with a reagent patch 1224 and hydrophobic valves 1223-1, 1223-2, and 1223-3 (collectively referred to herein as "hydrophobic valves 1223"). A hydrophilic mixing zone 1227 can be disposed between the longitudinal channel 1205-1 and the serpentine channel 1205-s to regulate the rate of sample flow between the sample pad 1211 and the test capture zone 1225. Additionally, in some embodiments, the hydrophilic mixing zones 1227 can be adjacent to the hydrophobic valve 1223-1, each having preselected dimensions (e.g., width and length) to achieve a desired fluid flow rate along the fluidic channel 1205. Die cut 1220 (or hydrophobic valve 1223) can be used to create a gated area that slows or restricts the flow of the assay fluid so that reagent patch 1224 can interact with the sample stream. The narrower the gate, the less sample flow there is. In some embodiments, slowing down one fluidic channel 1205 allows a slower fluidic channel to advance, so that all fluidic channels and all reagents arrive at test capture zone 1225-2 at more or less the same time, or nearly the same time.

[0060]

[0072] Embodiments consistent with test strip 1200 can include different combinations of longitudinal flow channels 1205-1 and serpentine flow channels 1205-s, along with hydrophobic gates 1223 and hydrophilic gates 1227, depending on the affinity of different reagents 1224 for their respective target analytes in the sample fluid. Thus, the selection of the shape and distribution of different flow components present in test strip 1200 can be varied according to the desire to obtain a rapid yet homogenous (e.g., near-simultaneous) response to different components of the assay in test capture zone 1225-2. In some embodiments, this is desirable to have a single endpoint for the assay test, thereby simplifying measurement and analysis logistics.

[0061]

[0073] Similar to test strip 1100 (see FIG. 10A), a first control capture zone 1225-1 at the beginning of fluidic channel 1205-s, proximal to sample pad 1211, and a second control capture zone 1225-3 at the end of the assay, closer to absorbent pad 1212, provide precise start and end points for the test. In some embodiments, a hydrophobic valve 1223-3 can be positioned in fluidic channel 1205-s to slow the end of the assay and ensure that test capture zone 1225-2 has fully interacted with the sample fluid before the end of the assay.

[0062]

[0074] Hydrophobic valve 1223, hydrophilic mixing zone 1227, and test capture zone 1225 are fluidic features contained in substrate 1201, shaped and sized to inhibit or facilitate sample flow across test strip 1200, as desired. In some embodiments, the details of the fluidic features of test strip 1200 are selected to provide time for a particular stage in the assay (e.g., for a reaction with one of reagents 1224 to occur or for conjugate immobilization in test capture zone 1225-2 to be completed). In some embodiments, the fluidic features of test strip 1200 can be selected to direct flow to distinct capture zones 1225 arranged in an array matrix. As disclosed herein, the capabilities of the fluidic features in test strip 1200 can be fully utilized by a digital capture device (e.g., as in image capture device 130, see FIG. 1).

[0063] How to use

[0075] 11 is a flowchart illustrating steps in a method 1300 of remotely diagnosing a disease using an image capture device, according to some embodiments. Method 1300 may be performed at least in part by a computer or image capture device, such as the architecture shown in FIG. 1. Accordingly, at least some of the steps of method 1300 may be performed by a processor executing instructions stored in a memory. Furthermore, methods consistent with the present disclosure may include at least one step as described in method 1300. In some embodiments, methods consistent with the present disclosure include one or more steps of method 1300 performed in a different order, simultaneously, approximately simultaneously, or overlapping in time.

[0064]

[0076] Step 1302 includes providing a device including a single, integral substrate, the substrate including multiple fluid flow channels and a single sample zone thereon that is common to each of the fluid flow channels, each fluid flow channel in direct fluid communication with the sample zone at a channel inlet region of the respective fluid flow channel. In the device, each fluid flow channel has a length and a width and includes a capture zone downstream from the channel inlet region and a channel constriction zone positioned between the channel inlet region and the capture zone, the channel constriction zone having a width and a length. The width of the channel constriction zone corresponds to a value within a range determined by (i) a minimum value equal to or greater than the diameter of a particular reagent deposited on the substrate, or (i') a minimum value equal to or greater than about 25% of the fluid flow channel width, and (ii) a maximum value equal to or less than about 75% of the fluid flow channel width.

[0065]

[0077] Step 1304 includes contacting the device with a biological sample from the subject.

[0066]

[0078] Step 1306 includes determining the presence or absence of a pathology or abnormality in the biological sample. In some embodiments, step 1306 includes determining the presence or absence of a bacterial infection, a viral infection, or drug addiction or misuse. In some embodiments, step 1306 includes determining the presence or absence of a viral infection, including a respiratory infection. In some embodiments, step 1306 includes determining the presence or absence of a bacterial infection, including Lyme disease or sepsis.

[0067]

[0079] Step 1308 includes diagnosing the condition or abnormality, if present in the biological sample.

[0068]

[0080] Step 1310 includes treating the condition or disorder with a suitable therapeutic agent. In some embodiments, step 1310 includes treating the condition or disorder with an antibiotic.

[0069] Hardware Overview

[0081] 12 is a block diagram illustrating an example computer system 1400 capable of implementing the image capture device and server of FIG. 1 and methods disclosed herein (e.g., method 1300, see FIG. 11), according to some embodiments. In some aspects, computer system 1400 can be implemented using hardware or a combination of software and hardware, in a dedicated server, integrated into another entity, or distributed across multiple entities.

[0070]

[0082] Computer system 1400 (e.g., server 110, image capture device 130) includes a bus 1408 or other communication mechanism for communicating information, and a processor 1402 coupled to bus 1408 for processing information. By way of example, computer system 1400 may be implemented with one or more processors. Processor 1402 may be a general-purpose microprocessor, a microcontroller, a digital signal processor (DSP), an application-specific integrated circuit (ASIC), a field-programmable gate array (FPGA), a programmable logic device (PLD), a controller, a state machine, gate logic, a discrete hardware component, or any other suitable entity capable of performing calculations or other manipulations on information.

[0071]

[0083] In addition to hardware, computer system 1400 may include code creating an execution environment for such computer programs, e.g., code constituting processor firmware, a protocol stack, a database management system, an operating system, or a combination of one or more thereof, stored in included memory 1404, such as random access memory (RAM), flash memory, read-only memory (ROM), programmable read-only memory (PROM), erasable PROM (EPROM), registers, a hard disk, a removable disk, a CD-ROM, a DVD, or any other suitable storage device, coupled to a bus, that stores information and instructions executed by processor 1402. Processor 1402 and memory 1404 can be supplemented by, or incorporated in, special purpose logic circuitry.

[0072]

[0084] The instructions may be implemented in one or more modules of computer program instructions stored in memory 1404 and encoded on a computer-readable medium to be executed by or to control the operation of computer system 1400, i.e., one or more modules of computer program instructions according to any method known to those skilled in the art, including computer languages such as, but not limited to, data-oriented languages (e.g., SQL, dBase), system languages (e.g., C, Objective-C, C++, Assembly), architecture languages (e.g., Java, .NET), and application languages (e.g., PHP, Ruby, Perl, Python). The instructions may also be implemented in a computer language such as an array language, an aspect-oriented language, an assembly language, an authoring language, a command line interface language, a compiled language, a concurrent language, a brace language, a data flow language, a data structure language, a declarative language, an esoteric language, an extensible language, a fourth generation language, a functional language, an interactive language, an interpreted language, an iterative language, a list-based language, a mini-language, a logic language, a machine language, a macro language, a metaprogramming language, a multi-paradigm language, numerical analysis, a non-English based language, an object-oriented class-based language, an object-oriented prototype-based language, an offside rules language, a procedural language, a reflective language, a rule-based language, a scripting language, a stack-based language, a synchronous language, a syntax processing language, a visual language, a wirth language, and an xml-based language. The memory may also be used to store temporary variables or other intermediate information during execution of instructions by the processor.

[0073]

[0085] A computer program as discussed herein does not necessarily correspond to a file in a file system. A program may be stored as part of a file that holds other programs or data (e.g., one or more scripts stored in a markup language document), in a single file dedicated to the program, or in multiple cooperating files (e.g., files storing one or more modules, subprograms, or portions of code). A computer program may be deployed to be executed on one computer or on multiple computers located at a single site or distributed across multiple sites and interconnected by a communications network. The processes and logic flows described herein may be executed by one or more programmable processors 1402 that execute one or more computer programs to perform functions by operating on input data and generating output.

[0074]

[0086] The computer system 1400 further includes a data storage device 1406, such as a magnetic or optical disk, coupled to the bus for storing information and instructions. The computer system 1400 can be coupled to various devices via an input / output module 1410. The input / output module 1410 can be any input / output module. Exemplary input / output modules include data ports, such as USB ports. The input / output module 1410 can be configured to connect to a communications module. Exemplary communications modules include networking interface cards, such as Ethernet cards and modems. In some aspects, the input / output module 1410 can be configured to connect to multiple devices, such as input devices 1414 and / or output devices 1416. Exemplary input devices 1414 include a keyboard and a pointing device, such as a mouse or trackball, by which a user can provide input to the computer system. Other types of input devices 1414, such as tactile input devices, visual input devices, voice input devices, or brain-computer interface devices, can be used to provide user interaction as well. For example, feedback provided to the user may be any form of sensory feedback, such as, for example, visual feedback, auditory feedback, or tactile feedback, and input from the user may be received in any form, such as acoustic, speech, tactile, or electroencephalographic input, etc. Exemplary output devices 1416 include display devices, such as an LCD (liquid crystal display) monitor, that display information to the user.

[0075]

[0087] In some embodiments, computer system 1400 is a network-based, voice-activated device accessed by a user. Input device 1414 and output device 1416 may include a microphone for providing queries in audible form in the user's language and for receiving inputs from the user, also in audible form. Additionally, in some embodiments, a neuro-linguistic algorithm may enable the voice-activated device to communicate with the user and receive the user's selection of a respiratory mask via a voice command or request.

[0076]

[0088] According to one aspect of the present disclosure, the image capture device 130 and the server 110 may be implemented using a computer system 1400 in response to the processor 1402 executing one or more sequences of one or more instructions contained in the memory 1404. Such instructions may be read into the memory 1404 from another machine-readable medium, such as the data storage device 1406. Executing the sequences of instructions contained in the main memory causes the processor 1402 to perform the process steps described herein. One or more processors in a multi-processing arrangement may also be employed to execute the sequences of instructions contained in the memory. In alternative aspects, hardwired circuitry may be used in place of or in combination with software instructions to implement various aspects of the present disclosure. Thus, aspects of the present disclosure are not limited to any specific combination of hardware circuitry and software.

[0077]

[0089] Various aspects of the subject matter described herein can be implemented in a computing system 1400 that includes a back-end component, such as a data server, or includes a middleware component, e.g., an application server, or includes a front-end component, such as an image capture device 130 having a graphical user interface or web browser through which a user can interact with embodiments of the subject matter described herein, or any combination of one or more such back-end, middleware, or front-end components. The components of the system can be interconnected by any form or medium of digital data communication, such as a communications network. The communications network (e.g., network 150) can include, for example, any one or more of a LAN, a WAN, the Internet, etc. Furthermore, the communications network can include, for example, any one or more of a network topology, including, but not limited to, a bus network, a star network, a ring network, a mesh network, a star-bus network, a tree or hierarchical network, etc. The communications module can be, for example, a modem or an Ethernet card.

[0078]

[0090] Computer system 1400 can include an image capture device and a server, where the image capture device and server are generally remote from one another and typically interact via a communications network (e.g., image capture device 130, server 110, and network 150; see FIG. 1 ). The relationship between the image capture device and the server arises through computer programs running on the respective computers and having an image capture device-server relationship with one another. The computer system can be, for example, but not limited to, a desktop computer, a laptop computer, or a tablet computer. The computer system can also be incorporated into another device, for example, but not limited to, a mobile phone, a PDA, a mobile audio player, a global positioning system (GPS) receiver, a video game console, and / or a television set-top box.

[0079]

[0091] As used herein, the terms "machine-readable storage medium" or "computer-readable medium" refer to any medium or media that participate in providing instructions to a processor for execution. Such media can take many forms, including, but not limited to, non-volatile media, volatile media, and transmission media. Non-volatile media include, for example, optical or magnetic disks, such as data storage devices. Volatile media include, for example, dynamic memory, such as memory. Transmission media include coaxial cables, copper wire, and fiber optics, including the wires that comprise a bus. Common forms of machine-readable media include, for example, floppy disks, flexible disks, hard disks, magnetic tape, any other magnetic media, CD-ROMs, DVDs, any other optical media, punch cards, paper tape, any other physical media with a pattern of holes, RAM, PROM, EPROM, FLASH EPROM, any other memory chip or cartridge, or any other medium from which a computer can read. A machine-readable storage medium can be a machine-readable storage device, a machine-readable storage substrate, a memory device, a composition of matter providing a machine-readable propagated signal, or a combination of one or more of these.

[0080]

[0092] As used herein, the phrase "at least one" preceding a list of items, with the term "and" or "or" separating any of the items, modifies the list as a whole, rather than each member of the list (e.g., each item). The phrase "at least one" does not require the selection of at least one item; rather, the phrase allows for the inclusion of at least one of any of the items, and / or at least one of any combination of the items, and / or at least one of each of the items. By way of example, the phrases "at least one of A, B, and C" or "at least one of A, B, or C" refer to A only, B only, or C only, any combination of A, B, and C, and / or at least one of each of A, B, and C, respectively.

[0081]

[0093] To the extent that terms like "comprising," "having," and the like are used in the specification or claims, such terms are intended to be inclusive, similar to the term "comprising" when interpreted as a transitional term in a claim. The word "exemplary" is used herein to mean "serving as an example, instance, or illustration." Any embodiment described herein as "exemplary" is not necessarily to be construed as preferred or advantageous over other embodiments.

[0082]

[0094] Reference to an element in the singular is intended to mean "one or more," not "one and only one," unless specifically stated otherwise. All structural and functional equivalents to the elements of the various configurations described throughout this disclosure that are known or later become known to those of ordinary skill in the art are expressly incorporated herein by reference and are intended to be encompassed by the technology of this application. Furthermore, nothing disclosed herein is intended to be made available to the public, whether or not such disclosure is expressly set forth in the description above.

[0083]

[0095] While the specification contains many specific details, these should not be construed as limitations on the scope of what may be claimed, but rather as descriptions of particular implementations of the subject matter. Certain features that are described herein in the context of separate embodiments can also be implemented in combination in a single embodiment. Conversely, various features that are described in the context of a single embodiment can also be implemented in multiple embodiments separately or in any suitable subcombination. Furthermore, while features may be described above as acting in certain combinations, and even initially claimed as such, one or more features from a claimed combination can, in some cases, be removed from the combination, and the claimed combination may be directed to subcombinations or variations of the subcombinations.

[0084]

[0096] Although the subject matter herein has been described with respect to particular aspects, other aspects may be implemented and are within the scope of the following claims. For example, while acts are depicted in the figures in a particular order, this should not be understood as requiring that such acts be performed in the particular order or sequential order shown, or that all of the depicted acts be performed, to achieve desirable results. Acts recited in the claims may be performed in a different order and still achieve desirable results. As one example, the processes depicted in the accompanying figures do not necessarily require the particular order or sequential order shown to achieve desirable results. In some situations, multitasking and parallel processing may be advantageous. Furthermore, the separation of various system components in the above-described aspects should not be understood as requiring such separation in all aspects, and it should be understood that the described program components and systems may generally be integrated together in a single software product or packaged in multiple software products. Other variations are within the scope of the following claims.

[0085]

[0097] In one aspect, a method may be an operation, an instruction, or a function, and vice versa. In one aspect, a claim may be modified to include some or all of the words (e.g., instructions, operations, functions, or components), one or more words, one or more sentences, one or more phrases, one or more paragraphs, and / or one or more claims recited in one or more other claims.

[0086]

[0098] To illustrate the interchangeability of hardware and software, various illustrative blocks, modules, components, methods, operations, instructions, algorithms, etc. have been described generally in terms of their functionality. Whether such functionality is implemented as hardware, software, or a combination of hardware and software depends on the particular application and design constraints imposed on the overall system. Those skilled in the art may implement the described functionality in different ways for each given application.

[0087]

[0099] As used herein, the phrase "at least one" preceding a list of items, with the term "and" or "or" separating any of the items, modifies the list as a whole, rather than each member of the list (e.g., each item). The phrase "at least one" does not require the selection of at least one item; rather, the phrase allows for the inclusion of at least one of any of the items, and / or at least one of any combination of the items, and / or at least one of each of the items. By way of example, the phrases "at least one of A, B, and C" or "at least one of A, B, or C" refer to A only, B only, or C only, any combination of A, B, and C, and / or at least one of each of A, B, and C, respectively.

[0088]

[0100] The word "exemplary" is used herein to mean "serving as an example, instance, or illustration." Any embodiment described herein as "exemplary" should not necessarily be construed as preferred or advantageous over other embodiments. Phrases such as "one aspect," "another aspect," "several aspects," "one or more aspects," "one embodiment," "an embodiment," "another embodiment," "several embodiments," "one or more embodiments," "one configuration," "configuration," "another configuration," "several configurations," "one or more configurations," the present technology, the disclosure, the present disclosure, and other variations thereof, are used for convenience and do not imply that the disclosure associated with such phrases is essential to the present technology or that such disclosures apply to all configurations of the present technology. Disclosure associated with such phrases may apply to all configurations, or to one or more configurations. Disclosure associated with such phrases may provide one or more examples. Phrases such as "one aspect" or "several aspects" may refer to one or more aspects, and vice versa, as applies to other phrases described above.

[0089]

[0101] Reference to an element in the singular is intended to mean "one or more," not "one and only one," unless otherwise stated. Masculine pronouns (e.g., his) include feminine and neuter genders (e.g., her and its), and vice versa. The term "some" means one or more. Underlined and / or italicized headings and subheadings are used for convenience only and do not limit the technology herein, and are not to be construed in connection with the interpretation of the description of the technology herein. Relative terms such as "first" and "second" may be used to distinguish one entity or operation from another without necessarily requiring or implying any actual relationship or order between such entities or operations. All structural and functional equivalents to the elements of the various configurations described throughout this disclosure that are known or later become known to those skilled in the art are expressly incorporated herein by reference and are intended to be encompassed by the technology herein. Furthermore, nothing disclosed herein is intended to be made available to the public, whether or not such disclosure is expressly set forth in the description above. No claim element is to be construed under the provisions of 35 U.S.C. 112, sixth paragraph, unless the element is expressly recited using the phrase "means for," or, in the case of a method claim, the phrase "step for."

[0090]

[0102] While the specification contains many specific details, these should not be construed as limitations on the scope of what may be claimed, but rather as descriptions of particular implementations of the subject matter. Certain features that are described herein in the context of separate embodiments can also be implemented in combination in a single embodiment. Conversely, various features that are described in the context of a single embodiment can also be implemented in multiple embodiments separately or in any suitable subcombination. Furthermore, while features may be described above as acting in certain combinations, or even initially as such, one or more features from a described combination can, in some cases, be removed from the combination, and a described combination may be directed to a subcombination or variation of the subcombination.

[0091]

[0103] Although the subject matter herein has been described with respect to particular aspects, other aspects may be implemented and are within the scope of the following claims. For example, while acts are depicted in the figures in a particular order, this should not be understood as requiring that such acts be performed in the particular order or sequential order shown, or that all of the depicted acts be performed, to achieve desirable results. Acts recited in the claims may be performed in a different order and still achieve desirable results. As one example, the processes depicted in the accompanying figures do not necessarily require the particular order or sequential order shown to achieve desirable results. In some situations, multitasking and parallel processing may be advantageous. Furthermore, the separation of various system components in the above-described aspects should not be understood as requiring such separation in all aspects, and it should be understood that the described program components and systems may generally be integrated together in a single software product or packaged in multiple software products.

[0092]

[0104] The title, background, brief description of the drawings, abstract, and drawings are incorporated herein into this disclosure and are provided as illustrative examples of the disclosure, not as limiting descriptions. It is submitted with the understanding that they are not used to limit the scope or meaning of the claims. Additionally, in the detailed description, the description provides illustrative examples, and it will be appreciated that various features are grouped together in various embodiments for the purpose of streamlining the disclosure. This method of disclosure should not be interpreted as reflecting an intention that the described subject matter requires more features than are expressly recited in each claim. Rather, as the claims reflect, inventive subject matter lies in less than all features of a single disclosed structure or operation. The claims are incorporated herein into the detailed description, with each claim standing on its own as separately described subject matter.

[0093]

[0105] The claims are not intended to be limited to the embodiments described herein, but are intended to be accorded full scope consistent with the language of the claims and to encompass all legal equivalents. Nonetheless, none of the claims are intended, nor should they be interpreted, to encompass subject matter that does not comply with applicable patent law requirements.

Claims

1. A plurality of separate fluid flow channels, including n fluid flow channels, where n is 2 to 20; a single, integral substrate comprising the plurality of separate fluid flow channels; a single sample zone on the substrate that is common to each fluid flow channel, each fluid flow channel being in direct fluid communication with the sample zone at a channel inlet region of each fluid flow channel; a capture zone downstream from the channel entrance region; a barrier extension region positioned in the channel entrance region; Equipped with each fluid flow channel having a length and a width, each fluid flow channel including a channel constriction zone positioned at the channel inlet region or between the channel inlet region and the capture zone, the channel constriction zone having a width and a length, the channel constriction zone width corresponding to a value within a range determined by (i) a minimum value equal to or greater than a diameter of a particle reagent deposited or to be deposited on the substrate, and (ii) a maximum value equal to or less than about 75% of the fluid flow channel width; each fluid flow channel n is identified by an integer from 1 to n, and fluid flow channel l and fluid flow channel n each comprise an outer channel wall having a width w, and the width of the barrier expansion region is from about w to about 10w; device.

2. 2. The device of claim 1, wherein the channel constriction zone length corresponds to a value within a range determined by (i) a minimum value equal to or greater than about 8% of the fluid flow channel length, and (ii) a maximum value equal to or less than about 75% of the fluid flow channel length.

3. 2. The device of claim 1, wherein the channel constriction zone length corresponds to a value within a range determined by (i) a minimum value equal to or greater than about 10% of the fluid flow channel length, and (ii) a maximum value equal to or less than about 65% of the fluid flow channel length.

4. The device of any one of claims 1 to 3, wherein the channel narrowing zone width is variable along the channel narrowing zone length.

5. The device of claim 4 , wherein the channel constriction zone comprises a tapered region at an entrance region to the channel constriction zone or at an exit region of the channel constriction zone.

6. The device of claim 5 , wherein the tapered region extends from the fluid flow channel width to the channel narrowing zone width.

7. The device of any one of claims 1 to 6, wherein the channel constriction zone is non-angled along its length.

8. The device according to any one of claims 1 to 3, wherein the particle reagent is an optically detectable solid particle.

9. 9. The device of claim 8, wherein the solid particles are fluorescent particles having a diameter of about 0.05 to 750 microns (0.00005 to 0.75 mm).

10. 9. The device of claim 8, wherein the solid particles are fluorescent particles having a diameter of about 0.05 to 10 microns (0.00005 to 0.01 mm).

11. 11. The device of claim 1, further comprising an entrance constriction region in each fluid flow channel, said entrance constriction region being positioned between said common sample zone and said channel constriction zone.

12. The device of claim 11 , wherein the entrance constriction region is positioned at the channel entrance region.

13. 12. The device of claim 11, wherein the entrance constriction region has a width and a length, the entrance constriction region width being substantially the same as the channel constriction zone.

14. The device of claim 11 , wherein the entrance constriction region is angled.

15. the inlet constriction region includes a bulge that bulges into the fluid flow channel; The device of claim 14, wherein the angle between the side walls of the bulge is about 30 to 90 degrees.

16. 16. The device of claim 15, wherein the bulge has a geometric shape that is a half diamond, a half rectangle, a half square, a quarter square, a quarter rectangle, a half parallelogram, a quarter parallelogram, or a half kite.

17. The device of claim 11 , wherein the entrance constriction region is non-angled.

18. The device of claim 1, wherein the barrier extension regions (422, 424) have a width of about w to about 5w.

19. The device of any one of claims 1 to 18, wherein the substrate is nitrocellulose.

20. A device according to any preceding claim, wherein each capture zone comprises a different capture reagent.

21. 21. The device of claim 20, wherein each capture zone comprises a capture reagent for an infectious agent.

22. 22. The device of claim 21, wherein the infectious agent is selected from respiratory syncytial virus, influenza A virus, influenza B virus, and human metapneumovirus.

23. 22. The device of claim 21, wherein the infectious agent is a Borrelia species.

24. 21. The device of claim 20, wherein each capture zone contains a capture reagent for a drug of abuse.

25. 25. The device of claim 24, wherein the drug of abuse is selected from fentanyl, buprenorphine, oxycodone, and 7-aminoclonazepam.

26. 21. The device of claim 20, wherein each capture zone comprises a capture reagent that distinguishes bacteria from viral infections.

27. 27. The device of claim 26, wherein the capture reagent comprises a reagent that binds to or interacts with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), C-reactive protein (CRP), interferon gamma-inducible protein-10 (IP-10), radical S-adenosylmethionine domain-containing 2 (RSAD2), MX dynamin-like GTPase 1 (MX1 or MxA), MX dynamin-like GTPase 2 (MX2 or MxB), neutrophil gelatinase-binding lipocalin (NGAL), and procalcitonin (PCT).

28. 28. The device of any one of claims 20 to 27, wherein the capture reagent is (i) a monoclonal or polyclonal antibody, (ii) a fragment of TRAIL, CRP, IL-10, RSAD2, MX1, MX2, NGAL, PCT, or (iii) a fragment of an infectious agent.

29. The device of any one of claims 1 to 28, wherein the substrate is a laminate comprising a hydrophobic material.

Images